Health Education: SPECIMEN COLLECTION FOR PATHOLOGY LABORATORY

SPECIMEN COLLECTION

Sample: Clinical specimens provided by the customer for testing. These include blood, serum, plasma, sputum, bronchial or gastric lavage fluid, urine, stool, cerebrospinal, pleural or ascitic fluid, tissue, etc.

PATIENT PREPARATION: - Many tests require that the patient be prepared in some specific way to ensure useful results. The best analytical techniques provide results that are only as good as the specimen that has been submitted for analysis. Our goal is to provide you with the most useful diagnostic information possible.

FASTING REQUIRMENT: - For the majority of test performed on serum, plasma or, whole blood, a fasting morning specimen is preferred.

The fasting specimen provided information that reflects the physiological baseline of the patient. This information can easily be compared to information from tests obtained at other times at provided a means for reliably monitoring a patients condition for the durations of care. From a practical stand point non fasting specimens are often lipemic, containing high triglycerides from food, with can interfere with many analytical procedure.

Blood, Serum and Plasma Specimens: - Most blood specimen can be obtained using routine phlebotomy techniques with few exceptions. The use of a tourniquet can cause stress an is net recommended in some case. The patient’s posture, sitting, standing or lying down, or the time of day relative to the patients sleep cycle can be important factors in some tests.

The three general procedures for obtaining are-

I. Venipuncture

II. Arterial puncture

III. Skin puncture

Blood is collection for various aspects of laboratory works-

I. Anticoagulated blood for hematology.

II. Serum for biochemistry, serology & blood bank.

III. Whole blood for blood culture.

IV. Plasma for coagulation studies.

Urine Specimen: - Many urine tests also require specific preparation of the patient for routine urinalysis the first morning voided (concentrated) specimen is always best for urine culture specimens, a mid stream specimen of urine should be submitted.

For urine Chemistry tests the 24 hr urine collection is the usual standard, for some of these tests there are directly restrictions that must be observed, for others, there are drugs that must be avoided prior to obtaining the specimen.

Proper Labeling of Specimens: - Each submitted specimen must be labeled with the patient’s name (written exactly as it appears on the requisition slip / test requisition form (TRF) and the test to be conducted. The tubes should be bar coded if possible.

Bar-coding Instruction:

1. Use appropriate barcode strickers.

2. No suffix strickers for biochemistry / hematology / immunology.

3. Affix stricker on center of the tube.

4. Number should be on the right hand side.

5. 90 sticker is used for blood sugar fasting and 99 for blood sugar post prandial. Do not use barcodes for other sugar sample.

Phlebotomist is explained the following basic steps for drawing a blood specimen

1. Check and confirm with the patient the details given on the Test Requisition Form.

2. Ascertain whether the patient is fasting and record the same on the Test Requisition Form.

3. Assemble the evacuated tubes, needle holder, alcohol swab, cotton swab, tourniquet etc. Required for phlebotomy. The phlebotomist should select the appropriate type of needle/ scalp vein set based on patient’s physical characteristics and amount of blood to be drawn. Keep the evacuated tubes in order of draw.

4. Reassuring the patient: The phlebotomist must gain the patient's confidence and assure him that, although the venipuncture is slightly painful, it would be of a short duration. Panic or anxiety of the patient will lead to difficulty in collecting the specimen.

5. Positioning the patient: The patient should be made to sit comfortably in a chair and should position his arm on a slanting armrest, extending the arm straight from the shoulder without bending at the elbow. If the patient wants to lie down, let the patient lie comfortably on the back. The patient should extend the arm straight from the shoulder. For support, a pillow may be placed under the arm. Check the prescription, test requisition form, evacuated tube type and their labels to ensure that all the details are matching and are appropriate for the tests ordered, before proceeding for collection.

6. For specimens collected for Immunohematology tests it is mandatory for the phlebotomist collecting the specimen to place his / her initials on the specimen tubes.

PROCEDURE FOR PHLEBOTOMY

Preparing for blood collection

1. Patient identification: - Check patient’s registered lab number on test request form with the bar code numbers to be applied on blood collection tubes.

2. Patient’s position: - Patient should be suitably positioned for venipuncture, either sitting or recumbent. This position should ideally be maintained for at least 15 minutes prior to performing the venipuncture.

3. Preparation for collection material: - Prior to venipuncture the following items must be prepared.

a). Blood collection system (Needle, Tube Holder & Vacutainer tube/s)

b). Bar code label/s for pasting on vacutainer tube/s

c). Sterile disposable gloves

d). Sterile swab

e). Disinfectant

f). Adhesive bandaid

g). Tourniquet

h). Disinfectant (Methylated spirit or 70 % Alcohol) swab.

i). A tray of cold water to rinse the needle & syring after drawing blood (by

syring method)

Note: - Tests requiring special collection conditions should be strictly adhered to prior to venipuncture.

BLOOD COLLECTION

1. Holding the green coloured section of the needle shield in one hand, twist and remove the white section with the other hand.

2. Screw needle on to the holder, leave the green coloured shield on the needle.

3. Prepare the venepuncture site remove the green coloured sections of the needle shield perform venepuncture in the usual manner with the arm in the downward position.

4. Introduce the appropriate vacutainer tube into the holder, placing your forefinger and middle finger on the flange of the holder and thumb on the bottom of the tube push the tube to the end of the holder puncturing the diaphragm of the stopper. Remove the tourniquet as soon as blood begins to flow into the tube.

5. When vacuum is exhausted, blood ceases to flow. Apply soft pressure with the thumb against the flange of the holder to disengage stopper from the needle and remove tube from the holder. If more sample are needles, repeat step 4.

6. Remove the last tube from the holder before withdrawing needle from the vein.

7. Gently invert all blood filled tubes 8-10 times to mix additives with blood. Do not shake. Vigorous mixing may case hemolysis.

8. Discard the used needle and insert a fresh one on the holder for the next patient. In case of accidental contamination of the holder, discard and replace with a new one.

Note: - Alternative draw blood in syringe; puncture the diaphragm of the appropriate Vacutainer tube/s with needle without removing the Colour-coded stopper. Allow to fill automatically till blood flow ceases. Do not force excess blood into tube.

Selection and Preparation of Vein Site for Blood Collection

Selecting vein site: For most venipuncture procedures on adults, veins located on the arm are used. The median cubital vein is most commonly used for the patient. If the venipuncture of this vein is unsuccessful, one of the cephalic or basilic veins may be used.

The following conditions should be avoided while selecting a vein: -

1. Vein from area having extensive scarring

2. Phlebotomy must not be performed on any size of hematoma.

3. Specimens should not be collected from the arms having intravenous site.

Procedure for vein selection

Locating veins

To locate veins it is necessary to palpate & trace the path of the veins several times with index finger.

Alternate site

Site such as dorsal wrist or hand & ankles or lower extremities may be required for patients with difficult veins.

Disinfection of the venipuncture site:

The puncture site must be cleansed to prevent microbiological contamination of the specimen and infection at the venipuncture site.

Cleansing is done with gloved hands:

Spirit or 70% ethanol is used for disinfection. A cotton ball is soaked in the spirit, excess should be dripped away. The cleaning should start from the vein and move out in a circular motion towards the outer surface. Allow the area to air dry to prevent hemolysis of the specimen, give enough contact time for the alcohol to bring out the disinfection of venipuncture site & prevent the patient from experiencing a burning sensation when the venipuncture is performed. Once disinfected, this site should not be touched with bare hands.

Applying the tourniquet:

A tourniquet is used to increase venous filling. This makes the vein more prominent & easier to enter.

Precautions when using a tourniquet:

The tourniquet should be released after no more than one minute. Local stasis can occur with hemoconcentration & the possible formation of a hemotoma due to infiltration of blood into tissue. This may result in erroneously high values for all protein based analytes, PCV & other cellular elements. If the test requested is Calcium then use of tourniquet is not permissible.

Tourniquet tying locations

Wrap the tourniquet around the arm, 3 to 4 inches above the venipuncture site. The tied tourniquet should never be left on the arm for more than 2 minutes because a tourniquet prevents blood from

flowing freely.

Collection Technique

Screw the needle into the holder. Puncture the selected vein at 30o to 40o angle. Insert the selected evacuated tube into the holder by pushing through the rubber sleeve of the rear cannula of the needle. After the vacuum in the tube draws the blood upto the mark on the tubes, withdraw the tube and proceed in a similar fashion if more evacuated tubes are

indicated. After blood has been drawn, the patient should release the fist & the tourniquet is also released and then the needle is withdrawn from the vein with simultaneous application of cotton at the venipuncture site.

The blood in the anticoagulated tubes is mixed by gently inverting the evacuated tube 7 to 8 times & blood collected in the plain (red top) tubes is kept at room temperature for 45 – 60 minutes for clotting and serum separation.

Note: Blood collected in plain tubes with clot activator additive is mixed by gently inverting the evacuated tube 4 to 5 times and then kept at room temperature for 30 – 45 minutes for clotting and serum separation.

Preventing hematoma during venipuncture

Puncture only the uppermost wall of the vein.
Remove the tourniquet before removing the needle.
Use the major veins.
Do not make partial penetration with needle.
Apply a small amount of pressure to the area with cotton after blood collection.

Collection of Blood from Pediatric Patients

Pediatric collections are done with the help of scalp vein needle (butterfly needle) and the blue top evacuated system needle (leur adapter). The end of the butterfly needle rubber tubing has to be fixed on to the blue top needle which is fitted on to the holder and used with the required evacuated tubes.

A cotton ball is held firmly over the venipuncture site as soon as the needle is removed. After checking that there is no blood flow a stick plast is applied at the site of venipuncture and the patient is given a cotton ball to be held at the site of venipuncture which has to be removed after 5-10 minutes.

In case of continued bleeding

Apply pressure to the site with a gauze pad until the bleeding stops.
Hold cotton firmly for sometime on the site & then put a bandage.
Tell patient to keep it for 15 minutes.

Disposal of Puncturing Unit

Dispose off needles promptly in a puncture resistant container with 1% sodium Hypochlorite.

Details of Specimens – Types, Collection & Processing

The pricelist available has tests listed in an alphabetical order with specific test methodology, specimen type and storage & transport conditions required for each test.

This pricelist is referred by the phlebotomist to obtain specific test related information before proceeding to test ordering & specimen collection. The specimen types are – Whole blood, serum or plasma (EDTA, Fluoridated, Citrated, Heparinised, Etc.), Urine, Body fluids, sputum, and stool.

Procedure for Specimen Collection

A. For whole blood

Blood is collected in a labeled (Patient's name, identity no.), specific anticoagulant containing evacuated tube (i.e. containing specific anticoagulant as per test specifications e.g. lavender top EDTA for CBC), to a full draw.
This specimen containing tube is gently inverted seven - eight times, to ensure proper mixing of blood and the anticoagulant. The specimen quality must be is checked, and should be free from hemolysis, clots and must be in recommended quantity.

B. For serum

Blood is collected in a labeled (Patient's name, identity no.) red top plain evacuated tube.
Allow the blood to clot for 45 -60 mins.
Centrifuge the specimen at 3500 rpm for 10 minutes.
Ensure that the information on the white cap plastic transfer vial and the evacuated tube from which the separated specimen is being transferred is matching.
Transfer separated serum to this vial with a plastic transfer pipette after ensuring that the serum is free from hemolysis and turbidity.
Ensure that the quantity of serum specimen is sufficient for all the tests requested.

C. For plasma

Blood is collected in a labeled (Patient's name, identity no.), specific anticoagulant containing evacuated tube as per test specifications.
Blood containing tube is gently inverted 7-8 times to ensure proper mixing of specimen and the anticoagulant.
The tube is centrifuged at 3500 rpm for 10-15 mins.
The white capped plastic transfer vial is used to collect the separated plasma. This vial must be labeled specifying anticoagulant in which blood was collected to obtain plasma and time of collection e.g. EDTA plasma, Fluoride plasma Fasting or Post Prandial.
After confirming that the details on the specimen containing evacuated tube and the transfer vial label are matching, the separated plasma is transferred to this vial with the help of a plastic transfer pipette.
The obtained plasma must be free from hemolysis and turbidity.
Ensure that the quantity of plasma specimen suffices specimen requirement of all the specific plasma requiring tests.

D. For body fluids

Body fluid collected by the treating physician is brought by the patient and is received by the laboratory as it is.
The specimen container must be examined for adequate labeling indicating Patient's name andspecimen type. The requisition form must be filled up appropriately.

E. For collection of Tissue Biopsy

For Histopathology / IHC:

Tissue Biopsy must be received in 10 % formalin in a leak proof container
Quantity of formalin added to preserve the tissue specimen, must be ten times the volume of tissue biopsy.

For Microbiology:

Tissue biopsy must be received in normal saline in a leak proof container.
Do not accept sample in formalin for microbiological testing.

F. For timed collections

For some hormonal studies, specimens are collected at specific timings e.g. Cortisol test. The blood specimens are collected at 8.00am and 4.00pm. The timing of blood collection has clinical significance and hence must be clearly specified on the evacuated tubes and

subsequently on the transfer vial. The serum/plasma must be separated at the earliest, stored and dispatched as specified in the price-list.

Whole Blood Specimen collection for Cytogenetics assay

The Specimen must be sterile for the above test. To ensure this, the whole blood specimen must be collected directly into the green top Sodium Heparin evacuated tube (sterile) aseptically.

Specimens for Cytogenetic Evaluation Collected at Sites other than a Collection Centre

Specimens for cytogenetic evaluation collected by the treating physician is received by the phlebotomist after checking for proper labeling and prescription and is forwarded as it is to the laboratory.

Note: Clinical History & family or provisional diagnosis is mandatory.

Specimen Collection for Products of Conception Test

Specimens for products of conception test collected by the treating physician is received by the phlebotomist after checking for proper labeling and prescription and is forwarded as it is to the laboratory.

Note: -

· Specimen for POC should be received in adequate normal saline and not in formalin.

· Fetus should never be accepted for the above tests as transportation of fetus is illegal.

· Specimen for POC should be transported in proper leak proof and puncture proof

containers.

ANTICOAGULATS

Anticoagulant: - Those chemical substances which prevent coagulation of blood are called anticoagulant.

A). Oxalate: - It is of three types.

I). Potassium Oxalate: - This is used at a concentration of 2 mg /ml of blood it

cause shrinks of RBC there for it is not used in PCV, ESR.

II). Ammonium Oxalate: - It is used at a concentration of 1mg / ml of blood it

Causes’ swelling of RBC there for it is not used in PCV & ESR.

III). Wintrobe’s Mixture ‘or’ Balanced Oxalate: - It balances the swelling effect

of ammonium oxalate & shrinking effect of potassium oxalate it is used in

the ratio 3:2 part.

Ammonium Oxalate -----------1.2 gm.

Potassium Oxalate--------------0.8gm

Distilled Water to make--------100 ml

It be comes 20mg/ml. 0.1ml of solution transfer in a dry clean vial let it for dry extremely high temperature should not be used because it activate the function of anticoagulant.

Action: - Oxalate combines with calcium present in blood to form calcium oxalate. It is soluble in nature.

Disadvantages: - Oxalate dose not preserve the morphology of the cell well it is toxic in nature.

B). Ethylene Diamine Tetra Acetic acid (EDTA): - It is two types.

I). Na2 EDTA (Disodium EDTA)

II). K2 EDTA (Dipotassium EDTA)

Commercial name of there salt ‘Versene’ & sequestrene’ respective.

Amount- 2mg/ml of blood.

Preparation- Dissolve 1gm EDTA in 100ml of D/W. Transfer 0.2 ml of 1% in to

Dry clean vials. Than dry it EDTA is a powerful calcium chelating agent.

Or,

Preparation of 1 % EDTA Solution:

· Weigh 1 gm EDTA in analytical balance or physical balance.

· Take a 100 ml volumetric flask and add 50-60 ml distilled water.

· Weighed powder is poured in the same flask mix well.

· After complete mixing add distilled water up to mark of 100 ml.

· Stopper the flask and mixed by inverting

· The put it in reagent bottle and label it 1 % EDTA solution by w / v

Preparations of EDTA vial:

Pipette out 0.2 ml 1% EDTA solution in different vials.
Keep the vials in to the Hot air oven at 100 ⁰C for 10 - 15 minutes or at 70 ⁰C for 3 – 4 hrs or 37 ⁰C for overnight.
After fix time EDTA powder appears because of evaporation of water and this is sufficient for 2 ml of blood.

Action: - It combines with calcium present in blood to from calcium action.

Advantage: - i). It gives good preservation of cell morphology.

ii). It preserve blood more then 6 hr.

iii). It inhibited the clumping action of platelet, so it is a good anticoagulant

for platelet count.

Disadvantage: - It is slightly expressive then oxalate.

C). HEPARIN: - Heparin is a highly acid in nature and normaly present in body in small amount it is also known as biological anticoagulant. (Most cell secreat heparin)

Used- It is used in some hematology special test or for biochemical testing and electrolytes blood gases ‘or’ for blood used in transfusions (is open heart surgery)

Action: - It inhibits the action of thrombin on fibrinogen and also the formation of thromboplastin, heparin inhibits the activity of there enzyme and dose not destroy then its activity slowly wearer of and the blood will then clot.

-Heparin can be neutralized by potamine sulphate.

Disadvantage: - i). It is expensive.

ii). It is not satisfactory for smear since it is highly acidic gives blue

Coloration to the back ground.

iii). It prevents anticoagulation only for a limited line so heparinized

blood has to be processed immediately.

Table: Preparation of the anticoagulant vial

S.No.

Name of the anticoagulant

Douses

Action

Uses

EDTA: Disodium or Dipotassium salt of Ethylene Diamine Tetra Acetic acid

1 - 2 mg / ml of blood

● EDTA+ Ca⁺⁺ → Calcium acetate

Hb, TLC, DLC, ESR, PCV, RBC count, Platelet count and G6PD

▪ Double Oxalate: Wintrobe’s mixture.

▪ Ammonium and potassium oxalate 3:2 ratio

3 mg Ammonium oxalate + 2 mg potassium oxalate for 2.5 ml of blood

● Ammonium oxalate + Ca⁺⁺ → Ca⁺⁺ oxalate

● Potassium oxalate + Ca⁺⁺ → Ca⁺⁺ oxalate

-do- except G6PD

3.8 % Trisodium Citrate

0.4 ml / 1.6 ml of blood

(1:4)

● Sodium citrate + Ca⁺⁺ → Ca⁺⁺ citrate

ESR for Westergren’s method

3.2 % Trisodium

Citrate

0.2 ml / 1.8 ml of blood

(1:9)

● Sodium citrate + Ca⁺⁺ → Ca⁺⁺ citrate

PT, APTT (coagulation studies)

Sodium Fluoride and Potassium Oxalate

2 mg Na F and 2 mg potassium oxalate for 1 ml of blood

● Sodium Fluoride + Ca⁺⁺ → Ca⁺⁺ Fluoride

Blood Sugar, Urea estimation

Heparin

0.1 – 0.2 mg / 1ml of blood

● Destroy in thrombin & thromboplastin

L.E .Cell, Osmotic fragility test and Blood gases, Electrolytes

ACD: Acid citrate dextrose

15 ml / 100 ml of blood

(21 days)

● ACD + Ca⁺⁺ → Calcium citrate

Blood banking

CPD: Citrate phosphate dextrose

15 ml / 100 ml of blood

(27 days)

● CPD + Ca⁺⁺ → Calcium citrate

Blood banking

CPDA: Citrate phosphate dextrose adenine

15 ml / 100 ml of blood

(35 days)

● CPDA + Ca⁺⁺ → Calcium citrate

Blood banking

RELIABLE RESULT START WITH RELIABLE SAMPLE

Additional Blood Collection needle & accessories

1. Luer Adapter

2. Multi - Sample Needle.

3. Eclipse Needle

4. Safety – Lok

5. Blood Collections set

6. Standard Holder

7. Pronto Holder

________________________________________________________________________

Vacutainer Tube holder with tube Tube holder, tube & needle

Additional Blood Collection needle & accessories Stopper

________________________________________________________________________

TUBE GUIDE BLOOD COLLECTION WITH THE VACUTAINER SYSTEM

Colour coded vacutainer tubes are used according to the specimen type required namely serum, plasma or whole blood.

S.No.

Stopper Colour

Number of gentle inversions

Additives

Used for

Red

● No Additive

● Clot activator

● None

Collection of Serum

Lavender

● EDTA

● Liquid K3 EDTA

● Spray-dried K2 EDTA

Whole Blood Collection

Green

● Heparin

● Sodium heparin

● Lithium heparin

Inhibits thrombin activation

Blue

● Buffered Citrate

Coagulation Studies

Black

● Buffered Sodium Citrate (0.4

ml)

● 129M Sodium citrate (3.8%)

Westergren’s ESR

Grey

Glycolytic inhibitor for glucose

● Potassium oxalate/sodium

fluoride

● Sodium fluoride

● Lithium iodoacetate

● Lithium iodoacetate/Lithium

heparin

● Na2 EDTA/Sodium fluoride

Glucose determination

Yellow

● Acid Citrate Dextrose (ACD)

● SPS-Sodium

polyanetholesulfonate

● ACD [Acid citrate Dextrose

Additives

I-Solution A-22.0%g/L trisodium citrate, 8.og/L citric acid, 24.5g/L dextrose,

II-Solution B-13.2g/L trisodium citrate, 4.8g/L citric acid, 14.7g/L dextrose.

Preserves red cells

Gold

● Clot activator and gel for serum

separation

Serum Separation

Light Green

● Lithium heparin and gel for

plasma separation

Plasma Separation

10.

Orange

● Thrombin

Inhibits thrombin activation

11.

Royal Blue

● Sodium Heparin

● None

Plasma/Whole blood and without anticoagulant Serum

12.

Brown

● Sodium Heparin

Plasma/Whole blood / Inhibits thrombin activation

13.

Light Blue

● 105M Sodium Citrate (3.2%)

● 129M Sodium Citrate (3.8%)

Coagulation Studies

14.

Bactec bottle/ Green

Transport Media/

Heparin

Microbial Cultures

Draw blood in the Colour coded vacutainer tube. For serum or plasma draw about 2- 2.5 times the requested volume. For serum, allow the blood to clot for at least 30 min. and separate by centrifugation. For plasma, mix the blood with the anticoagulant by gently inverting the tube 8-10 times and separate by centrifugation.

Holding the coloured section of the needle shield in one hand, twist and remove the white section with the other hand.

Screw needle into holder. Leave coloured shield on needle.

Prepare Venipuncture site. Remove the coloured section of needle shield. Perform Venipuncture in the usual manner with the arm in the downward position.

Introduce the tube into the older. Placing your fore finger and middle finger on the flange of the holder and the thumb on the bottom of the holder puncturing the diaphragm of the stopper. Remove the tourniquet as soon as blood begins to flow into the tube.

When vacuum is exhausted and blood flow ceases apply a soft pressure with the thumb against the flange of the holder to disengage stopper from the needle and remove tube from holder. If more samples are needed repeat from step-4

While blood is flowing into succeeding tubes, gently invert previously filled tubes containing additives 8 to 10 times to mix additives with blood. Do not shake. Vigorous mixing may cause haemolysis. Remove last tube from the holder before with drawing needle from vein.

Discard the used needle as per the safety guidelines of your institution. Never reshield by hand. In case of an accidental contamination of the holder we recommend discarding and replacing with a new one.

SPECIMEN COLLECTION ACCORDING TO TEST

HAE MATOLOGY

Hb / TLC / DLC / RBC Count / AEC / ANC / ALC / HCT / MCV / MCH / MCHC / CBC

2 ml fresh blood is collection in an EDTA VIAL.

If sample to be stored up to 6 hrs, it can be at room temperature and if it is to be stored up to 24 hrs then store it at 4 ˚C

ESR: -1.6 ml of whole blood collected in 0.4 ml tri-sodium citrate (4 volumes of blood to 1 volume of 109 mmol/L tri-sodium citrate) or 2ml of whole blood collection in EDTA.

Peripheral Smear for Malarial Parasite: - Any of the following.

1. EDTA Blood is the preferred sample

2. Capillary Blood

3. Stained or unstained slides

Reticulocyte Count: - 2ml whole blood in EDTA vacutainer.

The count will tend to fall slightly after 6-8 hrs unless the blood is kept at 4 ˚C satisfactory counts may be made up to 24 hrs if the blood is allowed to stand (unstained) at 4 ˚C

Peripheral Blood Smear:

1. EDTA Blood.

2. Stained or unstained slides. If only slides are received, note down the latest CBC report of the patient along with the date of the report.

The clinical diagnosis / history should be noted along with history of recent blood transfusion if any. The smear can be made before the blood is added to the anticoagulant or from EDTA blood. Ideally the films should be made within 1-3 hours of blood collection. However, EDTA blood may be stored at 4 ˚C for up to 24 hrs.

Blood Group: - 2 ml blood collection in a plain vial is required for tube method. Grouping can be done using EDTA blood sample by the tile / slide method.

Bone Marrow aspiration: - Bone aspirate slide along with peripheral blood smear slide / whole blood in EDTA.

APTT / PT: - Collect 1.8 ml whole blood in 0.2 ml of 3.2% tri-sodium citrate as anticoagulant. Though no special preparation of the patient is required prior to sample collection by approved techniques, it is preferable that patients are not heavily exercised before blood collection. Fasting or only light non-fasting meals prior to blood collection provide sample with a desirable lower opacity. History of drug intake should be noted especially if the patient is on oral anticoagulants or heparin.

Whenever possible, venous samples should be collected without a pressure cuff, allowing the blood to enter the syringe by continuous free flow. If a tourniquet has to be applied, it should be for the shortest possible time i.e. less than 1min. This is because venous occlusion causes haemoconcentration, increase of fibrinolytic activity, platelet release and activation of some clotting factors.

Withdraw blood without undue venous stasis or frothing into a plastic syringe fitted with a short needle of 19 to 20 SWG. The venepuncture must be a clean one and, if there is any difficult, take a new syringe and needle and try another vein. Transfer the blood into Anticoagulated tube, after detaching the needle from the syringe. Do not delay mixing blood with anticoagulant. Avoid foam formation during mixing. To minimize the effect of contact activation, good quality plastic syringes should be used. If glass blood containers are used, they should be silicone coated.

Mix exactly nine parts of freshly collected blood with one part of tri-sodium citrate (0.11 mol/l, 3.2%) Centrifuge immediately for 15 minutes at 1500-3000 rpm (approximately 1500 g ) on a laboratory centrifuge to obtain platelet poor plasma and transfer the plasma into a clean test tube. Cap the test tubes to prevent deterioration of samples. Plasma must be tested preferably immediately. However if the specimen are held at 22-24 ˚C then may be tested within 2 hours and if the specimen is held at 2-4 ˚C then they may be tested within 3 hours.

G-6-PD: - 2 ml blood in EDTA vial.

Heparin should not be used as it interferes with the reaction.

Sample may give false normal result in a deficient subject if the Reticulocyte count is high, as Reticulocyte have a higher G6PD activity than adult red cells. This is of special importance if the test is carried out immediately after a hemolytic episode due to a drug (primaquine or any such) in a sensitive subject. Hence obtaining history of any episode of bleeding and drug intake is very important.

HbA1c or GHB: - Venous (EDTA or heparin anticoagulated) sample or capillary blood sample taken from a finger prick.

Sample store with anticoagulants other than EDTA or heparin must not be used.

The venous sample may be stored at 2-8 ˚C for up to four days. Do not use samples which have been stored for more than four days. Store samples as whole blood, not as hemolysates or packed red blood cells.

Approximately 10ul of blood is required to run the test.

Coomb’s Test: - DAT: 2 ml whole blood in EDTA vial.

IAT: 2ml whole blood is collected in a plain vial.

(Direct & Indirect Each – EDTA / Serum)

TAKE SPECIMEN HEAMOTOLOYG TEST-2

Complete Hemogram / Platelet count / 2 ml fresh blood is collection in an EDTA VIAL.
L.E. Cells: Blood with glass bid
Sickleing test: - Citrate plasma
Osmotic Fragility: - EDTA Blood
Foetal Hb: - EDTA Blood
Bone Marrow Aspiration & Cytology: - EDTA Blood
Microfilaria: - EDTA Blood
Hb Electrophoresis: - EDTA Blood
Fibrinogen: - Citrate
FDP: - EDTA
CRT (Clot Retraction Time): - EDTA
Ceruloplasmin: - Serum
Body fluid count (Cell count Type): - Serum

Whole Blood Specimen collection for Cytogenetics assay

The Specimen must be sterile for the above test. To ensure this, the whole blood specimen must be collected directly into the green top Sodium Heparin evacuated tube (sterile) aseptically.

Specimens for Cytogenetic Evaluation Collected at Sites other than a Collection Centre

Note: Clinical History & family or provisional diagnosis is mandatory.

Specimen Collection for Products of Conception Test

Note: -

· Specimen for POC should be received in adequate normal saline and not in formalin.

· Fetus should never be accepted for the above tests as transportation of fetus is illegal.

· Specimen for POC should be transported in proper leak proof and puncture proof

containers.

To obtain plasma for coagulation test

Blood is collected in a labeled (Patient's name, identity no.), blue top Sodium Citrate evacuated tube.
Blood containing tube is gently inverted seven-eight times to ensure proper mixing of specimen with the anticoagulant.
The evacuated tube is centrifuged at 4000 rpm for 10-15 minutes.
Label the plastic transfer vial specifying anticoagulant used to obtain plasma and time of collection, Ensure that the above details match with the details given on the label of the specimen containing evacuated tube.
Transfer separated plasma to this vial with the help of a plastic transfer pipette after ensuring that the plasma is free form hemolysis and turbidity.
This plasma must be frozen and stored in the freezer compartment of the refrigerator till the time of processing.

Note: Coagulation tests are very sensitive specialized tests, proper specimen collection, processing and transport under specified conditions is mandatory to obtain accurate reports. Clinical History & or provisional diagnosis along with a list of medication taken by the patient must be noted, especially if it is oral anticoagulant like warfarin or intravenous anticoagulant i.e. Heparin.

Instructions to patients on oral anticoagulant therapy for prothrombin time

Many medications can affect prothrombin time test results. The patient has to submit the details of all medications taken before undergoing this test. There is no need to discontinue medication unless advised by the clinician.

Prothrombin time is usually done at the same time of day each time, so test results can monitor the dosage of medication used to prevent blood clots.

BIOCHEMISTRY

For timed collections

subsequently on the transfer vial. The serum/plasma must be separated at the earliest, stored and dispatched as specified in the price-list.

Total Protein (TP): - 2 ml serum is the preferred sample.

A reference interval for plasma would be 0.3 g / dl higher due to the presence of fibrinogen in the plasma.
A fasting specimen is not required but may be desired to decrease lipemia
Tightly stoppered samples are stable for 24 hrs at room temperature for one week at 2-8 ˚C

For plasma Separation following anticoagulants, may be used.

EDTA- 2 mg / ml of Blood.
Citrate- 6 mg / ml of Blood.
Heparin- 200 IU / ml of Blood.
Oxalate- 3 mg / ml of Blood.
Sodium fluoride- 10 mg / ml of Blood.

Albumin (Alb): - 2 ml serum is the preferred sample.

Plasma obtained from blood collection in EDTA vials may also be used.
Overnight fasting is preferred, Serum & Plasma sample. Ship refrigerated or frozen.
2 ml (1ml min.) CSF sample. Ship refrigerated or frozen.

Bilirubin: - 2 ml (1ml min. or 0.4 ml is required) Serum from 1 SST. Wrap tube in aluminum foil to protect from light. Ship refrigerated or frozen. Stability room Frozen 2 hrs 1 week 2 weeks.

Eventhough Serum is recommended, Plasma from Blood treated with Heparin can also be used for this test.
Only freshly collected Human Serum samples should be used for the determination of Direct Bilirubin, since direct reacting bile pigments that may appear in serum under certain conditions are reported to be unstable.
Serum samples must be kept away from both artificial and Sun light during processing and storage.
If samples cannot be assayed immediately after collection, special care must be taken to store them in dark under refrigeration.
Direct Sun light may cause up to 50 % decrease in Bilirubin with in one hour.
Bilirubin in Serum or Plasma is stable for one week if kept in dark under refrigeration.
Wrap tube in aluminum foil to protect from light.

For Lipid Profile

Fasting period of 12-14 hrs.

Cholesterol: - Serum

Overnight fasting sample is preferred.
LDL / VLDL Cholesterol / Lipid Profile / Total Lipid all test perform serum sample should be suitable.
The sample should be collected from a fasting individual and must not be Haemolyzed. Cholesterol values are slightly affected by meals, but more importantly the lipemic serum interferes with the direct cholesterol test by causing turbidity in the final reaction.
Cholesterol in the serum is stable for 7 days when stored at 2-8 ˚C.

HDL: - Serum is the preferred sample in our lab.

Plasma may be used if collected in any of the following anticoagulants:

EDTA 10 mg / ml Blood
Heparin 200 IU / ml Blood
Plasma should be separated immediately from the cells.
The sample should be collected after a 12 hour fast. Patients who are unable or unwilling to fast for this long, are requested to keep a 9 hour fast.
HDL Cholesterol value is stable in serum for 24 hours at 2-8 ˚C and 30 days when stored at -20 ˚C

LDH: - Serum

It should be free from hemolysis.
Total LDH is reported to be stable in serum for 1-3 days at 2-8 ˚C
Freezing inactivates the liver isoenzyme.

Triglyceride (T.G): - Serum is used in our laboratory.

Plasma obtained from blood collected in EDTA can also be used although T.G Concentration in EDTA plasma is about 3% less than in serum. Citrate should not be used as anticoagulant as it lowers the lipid concentration even more.
When plasma is used, blood should be cooled in an ice-bag as soon as it is drawn and cells separated within 3 hrs. Store the plasma at 4 ˚C till the test is done.
The sample is collected after 12 hrs of fasting. If the patient is unable to fast for 12 hrs he should be asked to give the sample after 9 hrs of fasting.
Blood is collected in the sitting position after the patient has been sitting quietly for about five minutes. This is because when the patient reclines, extravascular water transfers to the vascular system and dilutes the nondiffusible plasma constituents. This decreases the T.G levels. There the position of the patient should be standardized. If it is necessary to use the recumbent position, this position should be used each time, to minimize postural change.
Avid prolonged venous occlusion before venepuncture as it leads to haemoconcentration and increased level. Hence the tourniquet should be removed in one or two minutes.
Serum or Plasma sample may be refrigerated for up to 7 days if not tested within 24 hrs. For longer storage, sample may be frozen at -20 ˚C for up to 3 months.

Alkaline Phosphatase (ALP): - 2 ml serum is the preferred sample.

Plasma obtained from blood collection in Heparin (200 IU / ml blood) vials may also be used.
EDTA & Citrate should not be used as anticoagulant.
Blood should be collected in a clean dry container.
Haemolyzed specimen should be avoided as it may falsely elevate results.
Serum / plasma should be separated immediately from cells.
Alkaline Phosphatase is stable for 4 days at 2-8 ˚C and several months when stored at -10 ˚C

Acid Phosphatase (ACP): - 2 ml serum is the preferred sample.

Acid Phosphatase (Prostatic Fraction) / Acid Phosphatase (Total Prostatic Fraction): Serum sample preferred.

ALT (SGPT) / AST (SGOT):

Serum is required. Avoid prolonged contact with separated red cells.
Various anticoagulants have adverse effects on the enzyme activity. Hence serum is the preferred specimen.
The optimal temperature for ALT is 37 ˚C. At higher temperature its activity is decreased.
At>50 ˚C heat denaturation occurs. ALT is stable at 6 ˚C for atleast 24 hrs and at room temperature for lesser periods. Test should be carried out on the same dry as far as possible as ALT/AST activity decrease progressively with time. For prolonged storage, the temperature must be -20 ˚C or lower.
Hemolysis is associated with the release of enzymes from RBC in to the serum hence hemolyzed specimens (hemoglobin of 500 mg/dl) falsely elevate ALT results.

Amylase: - Serum / Heparinised plasma / Urine.

α Amylase is reported to be stable in the sample for 5 days at 2-8 ˚C
Separate serum from clot as soon as possible.

Lipase: - 2 ml serum is the preferred sample.

Uric Acid: - Serum / Urine

The patient should be in a fasting state for serum uric acid.
Serum should be separated from the cells at the earliest possible (within 30 minutes).
Uric Acid is stable for 3 days at room temperature (below 25 ˚C ) and for 6 months when stored at -10 ˚C
Collection of 24 hrs urine specimens should be made in containers with NaOH (10 ml of 5 % w/v) NaOH) to prevent urate precipitation.

Creatinine: - Serum / 2ml Urine in a sterile container.

Separate serum with in 1 hour of collection.
Creatinine in serum is stable for 7 days in the refrigeration and indefinitely when frozen. Creatinine in urine is stable for 2 to 3 days at room temperature and 5 days refrigerated.
Creatinine determination in urine is usually carried out on a 24 hour urine sample.
Thymol as preservative should be used for a 24 hour urine sample.
Random urine sample may also used. The type of urine sample collected must be clearly mentioned.

Creatinine Clearance Test:

Blood & Urine

Creatinine Phosphokinase: - Serum

It can be stored for 1 day at 15-25 ˚C and for 1 week at 2-8 ˚C

Phosphorous: - Serum / Urine

Acidity a 24 hrs. urine with specimen with concentrated hydrochloric acid ( 1ml /lit of urine), The acidified urine is stable for one week at 2-8 ˚C

GGT: -Serum or Plasma. Overnight fasting preferred.

Stability: At lest 1 week between -20 ˚C and + 25 ˚C
Discard contamination specimens.

Iron / TIBC: - Serum

It should be free from hemolysis.
Iron is reported to be stable in serum for 7 days at 2-8 ˚C.

Calcium: - Serum

Samples should preferably be collected in a fasting state.
Overnight fasting preferred.
Serum should be separated from the sample as soon as possible.

Glucose: - Serum / Plasma / CSF / Urine

For Plasma, collect venous blood in tubes containing oxalate- fluoride (2+2 mg / ml of blood).
2 ml (1 ml min) plasma from Sodium Fluoride tube. Overnight fasting is essential.
2 ml (1 ml min.) plasma each for Fasting and Post Prandial specimens from sodium Fluoride tube. Collect fasting specimen. Ask patient to have a normal to heavy meal. Note the time patient finishes the meal. Collect PP sample exactly after 2 hours.
Serum should be separated from blood as soon as possible.
Mix well by repeated inversions.
2 ml (1 ml min.) Body fluid in a sterile plastic screw capped container.

a. For fasting glucose:

Fasting period of 10-12 hrs required. Collect first voided mid-stream morning urine for urine glucose. Blood is collected by on duty phlebotomist.

b. For Post Prandial glucose:

Blood and urine specimens are collected after 2 hrs of meal taken.

Glucose Tolerance Test (GTT)

Blood samples collected in fluoride vial and Urine samples.
2ml (1 ml min) Plasma from 1 Grey top (Sodium Fluoride) tube and 10 ml (5 ml min.)Aliquot of corresponding urine for each timed specimen. Collect fasting baseline plasma and urine specimen (0 hr.) Administer 75 gm or 100 gm Glucose orally Collect additional plasma and urine sample at 0.5, 1.0, 1.5, 2.0 & 2.5 hours. Note time drawn on each tube, urine vial and test request from. Specify amount of glucose administered on test request from Overnight fasting essential ship refrigerated.

Preparation of patient

Prior to reporting for the OGTT, the patient must be clearly told that he would have to spend two hours in the lab during the test. For diagnosing gestational diabetes, a 3 hr GTT is done while if the patient is getting an extended GTT done, if would take 5 hrs.
The patient should be on balanced diet for three days prior to this test. (Containing at least 150 gm / day of carbohydrates.)
He must be instructed to report to the laboratory empty stomach after fasting for 12 hrs. He can drink water only.
Patient should avoid drugs likely to influence the blood glucose levels (insulin, oralantidiabetic drugs) for at least two days prior to the test.
During the test the patient should not cat food, drink tea, coffee or alcohol. He should not smoke or do any vigorous exercise.
He must be made comfortable and be seated during the test.

Sample collection and Glucose load dose

Fasting venous blood sample is collected in a fluoride tube.
Urine sample is also obtained.
First test the collected fasting urine sample for glucose. If glucose is present then don’t perform GTT by giving glucose, instead take post prandial sample.
The patient is then asked to drink 75 gm of anhydrous glucose dissolved in 250 -300 ml of water. He should drink it slowly (to avoid vomiting) and finish it in about 5 minutes.
The blood and urine sample are collected at ½ hr. interval for the next 2 hrs. (Total 5 samples including the fasting sample) If patient is unable to give four urine samples collect at least two urine samples at the one hr. intervals.

For the diagnosis of Gestational Diabetes, the method followed is slightly different:

Screening:

Is done by Glucose Challenge Test (GCT). A 50 gm oral glucose load is given. If the one hour post load glucose is 140 mg / dl or higher, a complete 100 gm 3 hr GTT is recommended.

GTT in pregnancy:

It is a 3 hr test where after collecting the fasting sample, 100 gm of glucose is given dissolved in water. Samples are taken at 1 hr interval till 3 hrs. Total four samples are taken.

Take Sample according to Test

1. Serum is the preferred sample: - Blood NPN / Blood Urea / BUN / Aldolase / Ironitibc / Chloride / Bicarbonate / Electrolytes (Total) / CPK / CPK-MB / LFT / Protein Electrophoresis / PBI / Inorganic Phosphate / Transferrin / Ferritin / CEA / PSA / Magnesium / Copper / Ceruloplasmin Copper / Cysticercosis / Zinc / CA 125 (Ovarian) / CA 15.3 (Breast) / CA 19.9 (G.I Tract) / Catecholamine / Anticaprdiolupin

2. EDTA Blood is the preferred sample: - Ammonia / Beta Micro Globulin in EDTA blood / Trop .T EDTA Blood

3. Urine is the preferred sample: - Urinary VMA / Urinary 17-Ketos Teroids (24 HRS) / Urinary 17-Ketogenic Steroids / Phenyl Ketonuria (Pyruvic Acid) / 24 hrs Urinary Uric Acid, Calcium, Protein, Creatinine (Each) / Beta Micro Globulin in urine / Microalbumin in urine / Urinary Copper

IMMUNOCHEMISTRY

Serum (including serum collected in separator tubes) or plasma (Heparin only) may be used.
For optimal result, samples should be free of fibrin, red blood cells or other particulate matter.
Specimens with obvious microbial contamination should not be used.
Samples may be stored for up to 24 hours at 2-8 ˚C prior to being tested. If testing will be delayed more than 24 hours, the serum or plasma should be separated from the clot or red blood cells and stored frozen (-10 ˚C or colder) for up to 12 months.

IMMUNOLOGY

Immunoglobulin IGG, IGM, IGA (Each): - Serum is the preferred sample
Australia Antigen: - Serum is the preferred sample
Toxoplasma Antibody: - Serum is the preferred sample
Alpha Feto Protein: - Serum is the preferred sample
Anti DSDNA: - Serum is the preferred sample
ELISA Test for (TB) IGG: - Serum is the preferred sample
ELISA Test for (TB) IGM: - Serum is the preferred sample
ELISA Test for (TB) IGG + IGM: - Serum is the preferred sample
ELISA Test for (TB) IGA: - Serum is the preferred sample
IGE : - Serum is the preferred sample
HIV (ELISA) : - Serum is the preferred sample
Total TORCH Panel: - Serum is the preferred sample
Vitamin B¹² : - Serum is the preferred sample
Vitamin D3 : - Serum is the preferred sample
Vitamin C : - Serum is the preferred sample
TPHA
SACE : - Serum is the preferred sample
P-ANCA & C-ANCA (Each) : - Serum is the preferred sample
Anti Smooth Muscle Antibody : - Serum is the preferred sample
ACTH : - Serum is the preferred sample
Anti Mitochondrial Antibody : - Serum is the preferred sample
APO Lipoprotein A: - Serum is the preferred sample
APO Lipoprotein B: - Serum is the preferred sample
PCR KOCH’S : - EDTA sample is preferred
ADA : - Serum is the preferred sample
FOLIC ACID : - Serum is the preferred sample

SEROLOGY

Mantoux Test : - Patient
Widal Test : - Serum is the preferred sample
ASO Titer : - Serum is the preferred sample
CRP : - Serum is the preferred sample
VDRL : - Serum is the preferred sample
Latex Fixation Test : - Serum is the preferred sample
ANF (ELISA) : - Serum is the preferred sample
HLAB27 : - EDTA sample is preferred

HEPATITIS PROFILE

1. HB,AG : - Serum is the preferred sample

2. HB,AB : - Serum is the preferred sample

3. HB۪ AG :- Serum is the preferred sample

4. HB۪ AB : - Serum is the preferred sample

5. HB Core Antigen IGM : - Serum is the preferred sample

6. Hepatitis B Profile (1 to 5) : - Serum is the preferred sample

To obtain Plasma for HIV viral load test

Blood is collected in a labeled (Patient's name, identity no.), lavender top EDTA evacuated tube to a full draw (till blood flow stops on its own or volume of blood collected is as stated on the tube).
Blood containing tube is gently inverted seven-eight times to ensure proper mixing of specimen and the anticoagulant.
This tube is centrifuged at 3500 rpm for 10-15 minutes.
Label the plastic transfer vial specifying anticoagulant used to obtain the plasma specimen and time of collection. These details must match the details on the evacuated tube label.
Transfer separated plasma to this vial with the help of a plastic transfer pipette, after ensuring specimen is free from hemolysis and turbidity.
This plasma must be frozen and stored in the freezer compartment of the refrigerator until time of processing

TESTS PERFORMED IN COLLECTION ROOM

(1) MANTOUX TEST

REAGENTS

PPD

Tuberculin PPD 1 TU: - For all patients

Tuberculin PPD 5 / 10 TU: - For patient with specific prescription from doctor.

Storage and Stability

Tuberculin PPD is stable at 2⁰ – 8⁰ C till expiry date mentioned on vial.

PROCEDURE

The tuberculin tests are based on reactions to intradermal innoculation of tuberculins.

Mantoux test: This is the method of choice and performed by intracutaneous injection of

0.1 mL of PPD into flexor aspect of forearm.

The preferred site for injection is surface of forearm.
Disinfect the site of injection with 70% alcohol and allow to dry.
Disinfect the tuberculin PPD vial stopper with 70% alcohol, allow to dry and then draw 0.1 mL of PPD by using tuberculin syringe.
Use 26G needle to inject the PPD at the site which is already disinfected.
Inject the PPD intradermally to make the deposition wheal, in the diameter of 6 to 8 mm which will raise up to the point of needle.
Mark the area of injection with indicator.
Instruct the patient not to apply any cream, oil, saops, and powder on this injected area.
Read the result after 48 and 72 hours.

MICROBIOLOGY

Blood for Microbiological Testing

Blood is collected aseptically in a labeled (Patient's name, identity no.), Green top Heparin tube / Bactec bottle.

Blood containing tube is gently inverted seven-eight times to ensure proper mixing of specimen and the anticoagulant.

Urine Collection

- Single urine specimens (fasting, midstream) are used for qualitative tests, but for quantitative tests a 24 hours urine sample is necessary.

- The phlebotomist provides a sterile container for specimen collection, single urine sample as well as 24 hours urine specimen.

- The single urine specimen should be transported to the laboratory as early as possible.

However, for 24 hours urine specimen collection details refer to:

Add suitable preservative to the container to be used for 24hrs. Urine collection. Refer the chart given along for details regarding Test and preservative to be used. Instruct the patient about the preservative and restrict him from discarding the same.
Total urine output in 24 hrs. is measured and noted, specify 24 hrs. urine sample on the Requisition Form.

Note: For Creatinine Clearance Test, also specify height, weight & age of the patient on the Requisition Form.

Send a well mixed 20 mL aliquot of the total volume for analysis. The container must be properly sealed to avoid leakage & suitably labeled stating it is a 24 hrs. sample with volume. The sample should be dispatched for processing at the earliest.

24 HOURS URINE COLLECTION

(Preservative to be added as per the test requirement)

S.No.	TEST	PRESERVATIVE	QUANTITY
1	Urinary Urea	Boric Acid	10 Gms
2	Urinary Phosphorus	Boric Acid	10 Gms
3	Urinary Uric Acid	Sodium Carbonate	5 Gms
4	Urinary Calcium	6 N HCl	10 mL
5	Urinary Magnesium	6 N HCl	24 mL
6	17 OH Corticosteroids	6 N HCl	25 mL
7	17 Ketosteroids	6 N HCl	25 mL
8	Catecholamines	6 N HCl	25 mL
9	Metanephrines	6 N HCl	25 mL
10	Urinary Protein	Boric Acid	5 Gms
11	Urinary Cortisol	6 N HCl	25 mL
12	5 Hydroxyindoleaceticacid	6 N HCl	25 mL
13	Vanillylmandelic Acid	6 N HCl	25 mL

Interfering substances for above tests

Urine Free Catecholamine: Catecholamine-containing drugs, Alpha-methyldopa, Isoproterenol, Isoetharine, Methenamine mandelate & Labetalol

Urine Metanephrines: Catecholamine-containing drugs, Alpha-methyldopa, Phenylephrine, Terbutaline, Metaproterenol, Phenothiazine, Methylglucamine

Urine Vanillylmandelic Acid (VMA): Catecholamine-containing drugs, Levodopa & Nalidixic acid.

For all the above assays, it is best to avoid Fenfluramine (large doses), Rapid Clonidine

Withdrawal and Alcohol (1 week)

No Preservative to be added for following tests:

Urinary Sodium.
Urinary Potassium.
Urinary Chloride.
Urinary Creatinine.
Urinary Glucose.

URINE COLLECTION FOR MICROBIOLOGICAL TEST

Collect urine sample from patient in regular sterilized Urine Container.
Put the urine transfer device in the container so that it dips in to the urine.
Insert the urine tube (evacuated) in to the transfer device, so that the needle penetrates the tube cap. The vaccum suction causes the urine to flow automatically in to the tube.
Remove the tube from the transfer device.
When the tube has correctly filled, invert it several times.
Dispose the urine container & transfer device.
Transport the urine tube to the lab.

Note: While sample collection and transit, preservatives add value in sample transportation and prevent unwanted growth. A urine sample collection in Boric acid stabilizer has advantages over methods like refrigeration of specimen or Dip-inoculation method. Preservative like Boric Acid overcomes limitations as it offers bacteriostatic action against Bacteria, Fungi in urine.

General considerations

This procedure addresses instructions that must be communicated to physicians, nurses,

phlebotomy teams, etc.

1. Instructions for proper specimen collection for Microbiological testing

Collect specimen before administering antimicrobial agents when possible.
Collect specimen with as little contamination from indigenous microbiota as possible to ensure that the sample will be representative of the infected site.
Utilize appropriate collection devices. Use sterile and aseptic technique to collect specimens to prevent introduction of microorganisms during invasive procedures.
Clearly label the specimen container with the patient’s name and identification number and with the date and time of collection before specimen collection.
Collect an adequate amount of specimen. Inadequate amount of specimen may yield false negative results.
Identify the specimen source and/or specific site correctly so that proper culture media will be selected during processing in the laboratory.
If a specimen is to be collected through intact skin, cleanse the skin first. For example, use 70% alcohol followed by iodine solution (1 to 2% tincture of iodine or 10% solution of povidine-iodine). Prevent burns by tincture of iodine by removing excess after specimen has been collected.
Before collecting the specimen, consider the risk/benefit ratio of the collection procedure to the patient.
Collect specimens in sturdy, sterile, screw-cap, leak proof containers with lids that do not create an aerosol when opened.

2. Instructions for proper specimen transport.

(a) Transport all specimens to the laboratory promptly.

1. To ensure the survival and isolation of fastidous organisms and to prevent overgrowth by more hardy bacteria.

2. To shorten the duration of specimen contact with some local anaesthetics used in collection procedures that may have antimicrobial activity.

3. To provide a more accurate diagnosis of the infectious disease process.

(b) Alternative to prompt delivery

Refrigerate most specimens at 2 to 8 ⁰C.

Specimens that may harbor temperature-sensitive organisms such as Neisseria species should be left at room temperature.

prior request from the laboratory.

(d) For Stool specimens

1. For bacterial culture, mix stool with a transport medium (Amie’s transport medium orbuffered glycerol saline).

2. For parasitology examination, mix stool with preservative.

(e) Hold CSF specimens at room temperature for culture, specimens meant for

biochemical tests only, may be stored at 2-8 ⁰C for 2-3 hrs.

For collection of Tissue Biopsy

For Histopathology / IHC:

Tissue Biopsy must be received in 10 % formalin in a leak proof container
Quantity of formalin added to preserve the tissue specimen, must be ten times the volume of tissue biopsy.

For Microbiology:

Tissue biopsy must be received in normal saline in a leak proof container.
Do not accept sample in formalin for microbiological testing.

Instructions for proper specimen collection, handling, transportation & preparation for Cytological testing

(a) GYNAEC CYTOLOGY:

COLLECTION OF VAGINAL, CERVICAL AND ENDOMETRIAL SMEARS:

When gynaecological smears are to be collected patient is instructed not to take a douche or insert any contraceptive drug or any other drug into vagina. Patient should not undergo prior digital examination, because the use of lubricant interferes with the cytological picture. Smears should not be collected during the menstrual period.

The patient is made to lie in the lithotomy position on an examination table. The labia are

separated & a dry bivalve speculum inserted gently into vagina without use of lubricant. With a sterilized wooden swab stick wrapped with a thin cotton wool, an adequate amount of material from vaginal pool (posterior fornix) is collected & spread rapidly & evenly in a rotary motion onto a clean glass slide previously marked ‘vagina’. Slide is immediately dropped into a fixative bottle or smear is covered with spray fixative to avoid air-drying.

Exocervix smear is collected from the squamocolumnar junction of exocervix (external os) on previously marked slide as exocervix & spread rapidly & evenly and fixed immediately.

Endocervix smear is collected by gently introducing swab stick in endocervical canal & rotating it in a clockwise & or anticlockwise direction & material is spread on slide marked as endocervix rapidly & evenly and fixed immediately.

In case of endometrial aspiration smears, material is aspirated out from uterine cavity by means of a metal cannula or a polyethylene tube connected to a 10ml syringe. The cannula slips easily through the external os into the uterine cavity.

Suction is then applied to the syringe & material is aspirated. Syringe is again disconnected & once again suction procedure repeated. In order to be sure that endocervical material is not mixed with that of endometrial, the syringe is first disconnected from canula & then canula is withdrawn. Smears prepared & fixed immediately. If more than one smear is fixed in same bottle, slides are placed back to back. The bottle containing slides in fixative can be sent to lab, along with relevant information of patient viz. age, date of last menstrual period, provisional clinical diagnosis, if any, pathological reports, gynaecological complaints if any, filled in requisition form.

TRANSPORTATION:

Smears transported to the main laboratory by different method:

Glycerine mailing technique.
Spray fixatives
95% ethanol or methanol

Note: the smears should not be dried and fixed immediately.

(b) NON-GYNAEC CYTOLOGY:

NON –GYNAEC SPECIMENS ARE:

Serous fluids : Ascitic fluid, pleural fluid, pericardial fluid
Cerebrospinal fluid
Urine specimen
Sputum sample
Miscellaneous: Pus aspirate, FNAC aspirate etc.

COLLECTION:

As soon as the non-gynaec specimen is aspirated, it has to be collected in a sterile plain container and refrigerated till transportation.

TRANSPORTATION:

Various body fluid, urine, sputum etc. are transported in a cold storage to the main laboratory for processing.

FNAC smears should be spray fixed or fixed in a 95% ethanol or methanol and transported.

PREPARATION:

Cloudy or turbid fluid:

The fluid as soon as it is received in the laboratory is stirred briskly with an applicator stick.

This causes the cells suspended in it to be dispersed and also prevents coagulation. A representative amount of the fluid is centrifuged for 10 minutes at 3000 rpm. The tissue fragments and clot, if present in fluid, is removed and cell block is prepared.

The supernatant part is discarded. An applicator stick with thin tightly wound cotton swab is dipped in the sediment button or buffy coat of the sediment and gently rolled over the surface of the slide.

It is important that the smears are made thin so that the cells lie in a monolayer. Two smears are made for each specimens.

Sparsely cellular fluids:

Clear and sparsely cellular fluid, specimens as well as urine and cerebrospinal

fluid yield scant or no sediment after centrifugation. In such cases a ytocentrifugation is used.

The cytocentrifuge concentration the small number of cells suspended in fluids. It prevents cell loss and gives optimum results in such sparsely cellular fluids. Spinning forcefully at 1000 r.p.m. for 10 minutes sediments the cells from a suspension directly onto albuminized glass slides. The suspension medium is simultaneously absorbed by the blotter or filter card. The result is a monolayer of well-preserved, well-displayed cells grouped within an area of 6 mm diameter on previously albuminized glass slides. Centrifugal force also flattens the cells offering a more open nucleus and more visually interpretable nuclear details. 2-4 drops (optimum 3 drops) are used for cytocentrifugation.

Major objections to use of a cytocentrifuge include distortion of cellular morphology due to air drying artifacts. This can be overcome by immediately fixing the smears after cytocentrifugation.

Sputum Cytology:

Sputum Cytology is the most commonly employed technique for the detection of lung cancer as it is simple, non-invasive, non-traumatic method, which gives high accuracy under optimum conditions.

In our Laboratory, either unfixed or prefixed sputum sample is received.

Processing:

Pick and smear method:

The sputum sample received in laboratory is first removed in the petridish.

By means of an applicator stick, whitish, opaque or blood-tinged, suspicious parts of the specimen are picked up put onto clean, prealbuminized glass slides and the material is teased out using the applicator stick or pressed between two glass slides.

Preparation of very thick uneven smears must be avoided as they give coverslipping problems.

Fixing is done in 95% ethanol and staining is done by the PAP method or H & E method.

For collecting cervical smears, insert the cervical brush into the os of endocervix and rotate in clockwise and anti-clockwise direction to get the specimen from endocervix and exocervix. After collecting cervical specimen using cervical brush, the head of the brush is detached from the cervix brush and placed into the LiquiPrep Preservative vial (Collection vial). After the specimen is collected in the LiquiPrep Collecion vial. Patient information is written on the vial label and the specimen is sent ambient to the laboratory for processing and analysis.

Other Cytological specimens

Other Cytological specimens or smears received in the laboratory are Breast discharges, oral smears, Bronchial secretions, oesophageal washing, gastric washing and lavage duodenal drainage, colonic washing, rectal aspiration, etc. are collected directly and sent to the laboratory as soon as possible.

URINE

Timed Urine Samples

These are obtained at designated intervals starting from time zero, and are noted on each

subsequent container with time of collection.

Collection of urine specimens:

Single specimens’ urine are used for ward examinations and for most qualitative tests, but for quantitative analysis, 24 hrs specimens are employed, except in the case of diastase, where a random sample will suffice. It is most convenient to collect the specimen from one morning to the next.

Changes on keeping:

Major changes which urine may undergo on keeping are due to bacterial action. The most important effect of bacteria is on the urea, which is converted into ammonium carbonate. Such urines are unsuitable for determination of urea, ammonia, total nitrogen and pH (since ammonia may be lost to the air). Microorganisms, both bacteria and yeasts, may also act on glucose.

If the urine becomes alkaline, phosphates may be precipitated on standing, uric acid and urates are deposited since they are less soluble in cool urine.

N.B: - Before carrying out estimations, any deposit must be well mixed with the urine before sapling. Uric acid and urates can be redissolved by moderate warming, phosphates by adding a little acid.

Urine routine & microscopy:

The patient must be instructed to submit a clean catch midstream sample.
The first morning sample is always preferable.
For random urine sample collection, first clean the genitals with soap and water and collect the midstream sample into the container. Close the container with a tightly sealable cap.
During collection the initial portion of urine stream is allowed to escape while the midstream portion is collected.

Urine for pregnancy test:

Should be the first morning voided sample.

Urine collection for microbiological testing:

Specimen: First morning midstream urine is the ideal specimen for microbiological testing but in case of urgency random urine specimen can be collected.

Instruction to be given to the Patient:

Collect 50-mL of first, morning midstream urine specimen in a sterile leak-proof container.
Use clean catch technique for urine specimen collection (Clean the urethral surface, open the sterile container just prior to passing urine.
Let some urine pass and thereafter collect the midstream specimen directly into the contain.

Urine culture:

The first morning midstream sample should be collected in a sterile container after washing the vulva / end of penis with soap and water, rinsing with sterile water and wiping with a sterile towel.

24 hour urine sample:

The patient is given a container for collection of 24 urine. Depending on the test required to be done the preservative is added. The patient is instructed not to throw it. He should be told to discard the first morning sample and start collecting all the urine passed during the next 24 hours into the container. The last urine to be collected will be the first morning sample on the second day of collection. The patient can collect each urine sample in a smaller clear container and pour it gently into the 24 hour container provided.
The entire container may be sent to the lab or 10 ml urine may be sent in a sterile container after thoroughly mixing the entire contents and noting the volume of urine voided in 24 hours.
From this time onwards collect all subsequent urine samples in the container provided. Collection should be continued till same time the next day (for e.g. 7.00 am). This is the 24 hrs. Urine sample.

Preservatives for urine:

If urines have to be kept, it may be necessary to prevent some of the changes described above. The choice of preservatives is often influenced by the purpose for which the urine is being collected.

Hydrochloric Acid (HCl):

As a rule, the use of acid is quit satisfactory; 10 ml of concentrated HCl is adequate for a 24 hour specimen. If it is considered undesirable to issue bottle containing the concentrated, 50 ml of 2N acid can be used. Such urines are suitable in particular for the determination of urea, ammonia, total nitrogen and calcium, but not for uric acid.

(Or, Add appropriate volume of 50 % or 6 N HCl to the 24hrs container for stabilization –to maintain pH between 1.0 & 3.0)

Thymol:

Thymol is also used as a preservative, either a few crystals, or 5 ml of 10 % (w/v) solution in isopropanol. The latter has been found to be satisfactory for a wide range of substances, viz. sodium, potassium, chloride, bicarbonate, calcium, urea, ammonia, uric acid, amino acids etc. but not suitable for 17 –oxosteroids when the Zimmerman reaction is used.

Formalin:

3 to 4 drops of formalin for 100 ml of urine can be used. If more is added, it may give false positive result with reducing tests for glucose.

Chloroform (CHCl₃):

Chloroform is also a satisfactory preservative but it may interfere with some tests such as the detection of glucose, since it reduces Fehling’s solution. The chloroform can, however, be boiled off. Since it settles to the bottom, chloroform mixes with any deposit. Sufficient chloroform is used to give a saturated solution in the urine.

Toluene and light petroleum:

These from a thin layer on the surface which tends to contaminate pipettes.

To preserve urine for determination of ascorbic acid, either 10 % concentrated acetic acid or 5 % metaphosphoric acid is used.

All these preservatives only prevent further surface contamination with bacteria; they do not stop the growth of bacteria already introduced during the passage and collection of urine.

For urine/sputum

Urine/ sputum specimen must be collected in the sterile plastic container.
The container is adequately labeled indicating patient's name and specimen type.

Preservation: In case of delay in testing, urine must be stored at 2-8⁰C (lower cooling

compartment of the refrigerator). There is no agreed-upon length of time for refrigeration as a preservative, because this depends on the individual urine constituents.

SPUTUM

Early morning sample is preferred.
On waking up the patient must rinse his mouth with water. He must cough up sputum from his lungs, put it in a sterile container and close the lid immediately. If sputum is not brought out.

Sputum Collection:

Specimen: Early morning specimen in a sterile plastic container.

Instruction to be given to the Patient:

The mouth to be rinsed well by using water.
The sputum must be coughed up from the lungs or bronchi and placed in the sterile plastic container. The container to be recapped immediately and secured tightly.

Semen

1. Semen analysis:

The patient should be instructed to collect semen after three days and no longer than five days of sexual abstinence in a sterile plastic container.
He should evacuate his bladder before ejaculation. Semen should be obtained by masturbation, and if circumstances preclude such collection, special silastic condoms should be used for collection with intercourse.
Only specimens collected in the laboratory will be processed and the same should be done within one hour of collection.

2. SEMEN ANALYSIS

PROCEDURE

On arrival, the Phlebotomist determines the period of abstinence from the patient and records this on the Test Requisition Form. Sterile wide-mouthed container is handed over, after labeling with patient name, identity number and date.

The patient is informed that the semen sample will need to be collected by masturbation.
Patient must be asked to evacuate his bladder before ejaculation.
The patient must be instructed regarding the mode of sample collection into the wide mouthed jar. Adequate tissue paper is given to him so as to ensure that the jar is clean from outside when handed back to the staff.
The patient is led to a quiet, secluded room which offers privacy to the patient during the sample collection period.
When the patient is handing over the container, the staff must determine whether any sample was lost during collection due to spillage or other reasons.
In the event that the patient is unable to collect the sample by this method, the referring doctor must be contacted for discussing options for sample collection. These might include

a. The use of special Silastic condoms for semen collection during intercourse or

b. Semen collection following prostatic massage.

Instructions for Report Pick Up

The patient is given a specific time and day for report pick up. The turn-around-time mentioned in the price list must be referred for the same. No over commitment regarding the time of report pickup should be made.

SAMPLE STABILITY

S.No.	Test	Sample	Stability 2-8 ⁰C	Stability2-6 ⁰C
1.	Glucose	Fluoride Blood	Up to 6 days	Up to 10 days
2.	Urea	Serum or (Fluoride Plasma, Heparin zed Plasma)	Up to 3 days	Up to 7 days
3.	Creatinine	Serum, Heparin zed Plasma	Up to 2 days	Up to 7 days
4.	Uric Acid	Serum, Heparin zed Plasma	Up to 5 days	Up to 10 days
5.	Total Protein	Serum	Up to 7 days	Up to 2 weeks
6.	Albumin	Serum	Up to 7 days	Up to 2 weeks
7.	SGPT	Serum, Heparin zed Plasma	One days	Up to 3 days
8.	SGOT	Serum, Heparin zed Plasma	One days	Up to 3 days
9.	S.ALP	Serum, Heparin zed Plasma	One days	Up to 7 days
10.	Cholesterol	Serum, Heparin zed Plasma	Up to 7 days	Up to 2 weeks
11.	T.G	Serum, Heparin zed Plasma	Up to 3 days	Up to 7 days
12.	Sodium	Serum, Heparin zed Plasma	Up to 7 days	Up to 2 weeks
13.	Potassium	Serum, Heparin zed Plasma	Up to 7 days	Up to 2 weeks
14.	Bilirubin	Serum, Heparin zed Plasma	One day	Up to 3 days
15.	PSA	Serum	24 hours	0 ⁰C for 3 days

Normal Colour of Serum and Plasma a Pale yellow

HANDLING OF INFECTIOUS WASTE

Gloves must be worn at all times when handling infectious waste.
Infectious waste is segregated from other waste at the point of origin (in special garbage bins which are appropriately labeled).
Infectious waste is kept separate from other wastes. Enclosures or containers used for the containment of infectious waste is secured so as to deny access to unauthorised persons and is marked with prominent warning signs on, or adjacent to, the exterior of entry doors, gates or lids.
Infectious waste, (except for sharps) is contained in disposable plastic bags which are impervious to moisture and sufficiently sturdy to prevent damage under normal usage. The bags are securely tied so as to prevent leakage or expulsion of solid or liquid wastes during storage, handling or transport.
Infectious sharps are contained for disposal in leak proof rigid, puncture-resistant containers which are closed, precluding loss of contents.
All bins used for containment and disposal of infectious waste are colour coded as per local regulatory requirements and conspicuously labeled with the international biohazard symbol.
Infectious waste in bags or other disposable containers shall not be subjected to compaction by any compacting device and shall not be placed for storage or transport in a portable or mobile trash compactor.

DISPOSAL OF WASTE

All infectious materials are discarded in the biohazard container as per the local regulatory requirements. The non-infectious waste is discarded in the general waste bin. All the contents of the biohazard container is sent to the local facilities available for further treatment of biohazard waste for disposal of the same.

All needle are discarded in a puncture resistant container filled with 1% solution of Sodium Hypochlorite. These containers are also sent alongwith the infectious waste containers for appropriate disposal.

Infectious wastes are handed over to the infectious waste disposal agency for further treatment.

NOTE: Needles should not be recapped, bent, or sheared, and should be placed intact into sharps containers, A one-handed recapping technique should be used whenever the needle must be removed/ recapped.

Preparation of 1% Sodium Hypochlorite:

From commercially available stock solution prepare 1% Sodium Hypochlorite solution as per vendor’s instructions. Fresh solution should be prepared daily and stored in dark coloured container away from sunlight.

Procedure for Decontamination of Spills

Prepare a 1% Sodium Hypochlorite solution

In case of leaked out specimens / broken glass tubes contaminating the counter top or the floor the following should be done.

a. Cover the spill with absorbent tissue and pour 1 % of Sodium Hypochlorite over the tissue and let it soak for 15-20 minutes.

b. At the end of 20 minutes carefully remove the soaked tissue (wearing gloves) to the biohazard container for disposal.

c. If broken glass is present, do not try to remove materials (including broken glass) with hands. Always use a small dustbin and scoop the material into it using a plastic dustpan to collect the material. The collected material should then disposed into the puncture resistant container with 1% Sodium Hypochlorite solution. The dustpan should be soaked in a basin containing 1% Sodium Hypochlorite solution for at least 2 hours.

d. Remove the gloves and discard them into the biohazard container.

e. Rinse your hands with Sterilium. Alternatively soap and water can be used.

LABORATORY SAFETY METHODS

A safe working environment can be maintained by instituting bio-safety practices that also include careful measures for personal hygiene, for cleanliness in the work space, and for proper handling of biohazardous materials. An understanding of these necessitates an awareness of potential biohazards while performing laboratory duties, and will help to prevent accidents, injuries and infections.

United States Occupation Safety and Health Administration (OSHA) recommendations for handling Infectious materials:

All fluids from all clients should be considered infective and handled and transported appropriately.
There should be no distinction between HIV positive and HIV negative samples. (One obvious reason for this ‘universal’ label is undiagnosed HIV patient). It is much safer for all concerned if all clients were to be considered as positive, for all specimens would then be treated with the same level of caution.
Cerebrospinal fluid, synovial fluid, pleural fluid, peritoneal fluid, amniotic fluid, and any blood product are included in the universal precaution warning.
Specimens not included are: nasal secretions, faeces, sputum, sweat, tears, urine and vomitus.

Accidental Exposure and Occupational Risks

The primary work-related dangers are parenteral exposure through accidental needle sticks, cuts from contaminated equipment, exposure of mucous membranes and exposure of chapped or broken skin, wounds, and scratches to contaminated specimens.

Although many potential routes of exposure exist, it has been reported that up to 80% of all exposure occur as the result of needle sticks.

OSHA Recommendations for avoiding accidental exposure are listed below

Dispose off needles properly in puncture resistant containers.
Wear latex or vinyl gloves whenever handling or obtaining body specimens of any kind. This is cumbersome, uncomfortable, but extremely necessary. Gloves will not give much protection from a needle stick, but they will prevent any skin contamination from a specimen, thus preventing inoculation through cuts or abrasions.
When gloves are removed or changed, wash hands.
Wear an apron and leave it in the laboratory.
Aprons and gloves must be used in the Collection area when handling any equipment, pipettes, specimens, refrigerators, and freezers where samples are stored.

A clean, organized workspace is a safer workspace. The following measures will help increase safety in the laboratory:

Sodium hypochlorite, alcohol should be well labeled, in proper containers and stored properly (example: store in a separate cabinet below benchtop level).
Keep benchtop free from non-essential materials.
Materials should not be placed near the edges of counters or shelves, but rather organized as related to need.
Hazardous materials and reagents not in use should be safely stored out of the immediate work area.

Countertops or work areas should be cleaned with disinfectant before and after each workday.

Other prudent measures and safety practices that help reduce the risk of infection include,

immediate containment of spills and wiping of the contaminated surface using the disinfectants

Biohazard Waste and Occupational Risks

A separate puncture-resistant container for sharps – glass waste and needle must be used.
Liquid waste is decontaminated by autoclaving or by adding sufficient sodium hypochlorite before discarding.

Universal Precautions

The proper method of self-decontamination after performing work is to remove the lab coat first, then gloves, and finally to thoroughly wash hands with soap or disinfectant in the ‘clean’ sink. If hands are washed before removing the coat, the hands may become contaminated again while handling the coat.

The following practices are employed

Protective shoes that completely cover feet are worn (no open toe shoes or sandals).
Gloves are worn when handling blood or body fluids.

FIRST AID

Thrombophob ointment/icepack to be applied at site of hematoma.
In case the patient faints he/she has to be made to lie down & his feet have to be raised by placing pillows under his feet so that the blood flows to the brain.
In case the patient is feeling faint due to hypoglycemia (extended fasting hrs.) then give him glucose to overcome the same.
A physician located in the immediate surrounding area must be on call for any emergency & his name; contact number must be easily available to the phlebotomist.

CRITERIA FOR SPECIMEN REJECTION

Causes for specimen rejection:

Specimen received in formalin for culture. (If exposure to formalin is short i.e. < 24 hours) one may be able to culture certain organisms such as Mycobacteria if sample is collected from the innermost part of the specimens not exposed to formalin).
48 hour old sputum samples for culture.
Samples which have leaked out of the container.
Submission of samples in unsterile containers for bacterial culture.
Dried out swabs and culture plates that contain bacterial growth.
Specimens that are contaminated with barium (stool), coloured dyes or oily chemicals.
Samples that are difficult to recollect (CSF, bronchial washing etc) should be processed anyway. However, on the final report one should indicate the condition of the sample received to caution the doctor regarding the possibility of specimen contamination at collection or at transport. (i.e. report with a disclaimer).

ACCOUNTABILITY

Phlebotomists are responsible for implementation of the policies with respect to the preanalytical procedures.
GM-Operations/ Lab Heads are responsible for enforcement within laboratory.
Executive Director – Operations / Chief Quality Officer is responsible for enforcement of policy with respect to the entire laboratory.

REFERENCES

CLSI Infobase 2009, GP05-A2: Clinical Laboratory Waste Management; Approved Guideline-Second Edition.
Textbook of Medical Laboratory Technology: Praful B. Godkar.
Clinical Diagnosis & Management by Laboratory Methods: John Bernard Henry.
TIETZ Textbook of Clinical Chemistry and Molecular Diagnostics: Carl A. Burtis, Edward R.Ashwood, David E. Bruns; Fourth Edition.

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Health Education

Sunday, March 16, 2025

SPECIMEN COLLECTION FOR PATHOLOGY LABORATORY

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