INSTITUTE OF MANAGEMENT STUDY

AFFILIATED TO MAULANA ABUL KALAM AZAD UNIVERSITY OF TECHNOLOGY

INTERNSHIP PROJECT REPORT AT

KPC MEDICAL COLLEGE AND HOSPITAL
FOR FULFILMENT OF BACHELOR OF MEDICAL LABORATORY TECHNOLOGY

SUBMITTED BY
NAME ; MOHIMA YESMIN
ROLL NO ; 19441423012
REG NO ; 231942410021
DEPT OF B.M.L.T
 ACKNOWLEDGEMET
‘’Presentation , Inspiration , and Motivation have always played a key role in the
success of any venture’’

I express my deepest thanks to our Respected Director sir,

Mr. Tapash Ranjan Saha & Mrs. Rajrupa Ghosh [ Head of the Department]
of INSTITUTE OF MANAGEMENT STUDY , for his valuable advice about our duty and
responsibility as a technologist in the training period .

I also convey my sincere thanks to our class teachers and all

the honorary guest teachers for their inspiration and advice for the internship and report
preparation.
I would like to convey my sincere thanks to the technical staff of the
KPC MEDICAL COLLEGE AND HOSPITAL for their kind co- oporation and help throughout
the internship preiod .

I also convey my sincere thanks to the faculty members Mrs. Rajrupa

Ghosh [HOD of BMLT Dept] , Mrs Moumita Chakraborty, Mrs Moumita Sarkar mam , IMS
& all the honorary teachers for their inspiration & advice throughout the courses as well
as for the internship training and report preparation.

Finally I must express my very profound gratitude to my

parents and family members for providing me with unfailing support and continuous
encouragement throughout my years of study and during the internship preiod ,and this
report preparation . This accomplishment would not have been possible without them.

Thank You
MOHIMA YESMIN
 INTRODUCTION
The internship is an integral part of the program for Bachelor of
Medical Laboratory Technology and is designed to provide interns
with an opportunity to integrate and apply previously acquired
knowledge and technical skills in actual clinical settings. Under the
guidance of experienced Medical Laboratory Professionals and
other qualified laboratory personnel and health professionals,
interns learn more about diagnostic test procedures, quality control
methods and programs, and instrumentation in the clinical
laboratory. They also gain an understanding of the roles and
functions of the Medical Laboratory technologist. The internship
provides applied learning experiences during which the intern
should
a. Acquire clinical laboratory skills
b. Perform quality control procedures
c. Learn and adapt new procedures
d. Operate and maintain various laboratory machines and
instruments. Report accurate and precise results to supervisors
e). Understand the responsibilities, roles, and functions of the
Medical Laboratory technologist.

Internship Discipline And Duration

Biochemistry 04.08.2025 to 03.09.2025

Central laboratory 04.09.2025.2025 to 03.10.25
Sample collection 04.10.2025 to 3.11.2025
GENERAL LABORATORY SAFETY PROCEDURE AND RULES

ROLES OF LABORATORY TECHNOLOGIST

 Maintain the cleanliness and safety of the laboratory.

 Ensure that the equipment and glassware are kept safe and clean.

 Handle and maintain the microscope.

 Sterilize the equipmentes as per required.

 Dispose of specimens and infected materials in safe manner.

 Maintain the necessary reports of investigations done and submit the reports of
the superintend/BMOH/ MO. (as the case may be).

 They will make “Indent for materials and supplies” - for the laboratory through
the superintend/BMOH/ MO. (as the case may be) and ensure the safe storage of
materials and supplies received.

 Prepare “Periodical Reports” regarding his work and authenticate for

transmission.

 Practice the thumb rules of Bio-Medical Waste Disposal.

Personal Safety Procedures for Lab Technologists

. Wear PPE: Lab coats, gloves, masks, and eye protection

. Handle samples with care: Assume all samples are infectious

. Avoid touching face: Especially eyes, nose, and mouth

. Use pipette aids: No pipetting by mouth

. Work in well-ventilated areas: Avoid inhaling chemicals or biological

agents

. Wash hands frequently: Before and after handling samples, equipment,

and after removing gloves

. Shower and change clothes: After lab work, especially if contaminated

. Avoid eating or drinking: In the laboratory or near lab equipment

AIMS AND OBJECTIVE

To learn the advance techniques of independent approach, manners as well as

development of personal skill.

 To learn the principle operational technique to use different modern and

sophisticated instruments used for human health and health related disorders.

 To gain knowledge about the patient handling and how to give instruction to the
patient before the collection of samples

. To gain knowledge about collection, preservation, testing process and handling

of individual sample according to the disease condition of the patient

 To learn at the time of test how to handle different hazardous and non-hazardous
sample and after how to dispose such type of samples.
 :Research lab of BIOCHEMISTRY :
INTRODUCTION ;
The Biochemistry Research Laboratory is an important part of medical science where
different types of hormonal tests are performed to diagnose diseases. The laboratory
uses modern automated instruments such as the ADVIA machine, which provides
accurate and reliable results.

In this lab, various hormone tests are done, including TSH, FT4, FT3, T4, T3, FSH, PRL, PSA,
and Ferritin (FER). These tests help in detecting thyroid disorders, reproductive and
hormonal problems, prostate conditions, and disorders related to iron metabolism.

All the tests are performed under standard laboratory procedures, maintaining proper
quality control for accurate results. The Biochemistry Research Laboratory thus plays a
vital role in diagnosis, treatment, and research in the field of medical biochemistry..

PRINCIPLE OF BIOCHEMISTRY RESEARCH LAB ;

.It is based on the idea that changes in hormonal levels reflect normal or diseased
conditions in the body.
. Tests are performed using specific chemical reactions and antigen–antibody
interactions.
. Automated instruments (like ADVIA) measure test results through photometry,
chemiluminescence, or immunoassay principles.
. The intensity of the signal produced (color/light) is directly proportional to the
amount of the analyte in the sample
. Standard procedures and quality control are followed to ensure accurate and
reliable results.
INTRODUCTION AND PRINCIPLE OF ADVIA MACHINE
The ADVIA is a fully automated, high-performance chemiluminescent
immunoassay analyzer used in the Biochemistry Research Laboratory for
accurate and efficient hormone testing. It is designed to perform a wide range
of immunological tests with high precision and reliability.
The ADVIA system uses the Chemiluminescent Immunoassay (CLIA) principle
to measure various hormones, enzymes, and proteins present in human serum
or plasma. It automatically performs all steps — including sample
identification, reagent dispensing, incubation, washing, detection, and result
calculation — without manual interference.
In this technique, specific antibodies bind to the target hormone (antigen)
present in the sample. A chemiluminescent substrate is then added, which
reacts with the enzyme-labeled antibody to produce light. The amount of light
emitted is directly proportional to the concentration of the hormone in the
sample.
This advanced instrument provides fast, reproducible, and accurate results,
making it highly useful for clinical diagnosis and research. The ADVIA machine
is widely used for hormone assays such as TSH, FT4, FT3, T4, T3, FSH, PRL,
PSA, and Ferritin, helping in the detection of thyroid, reproductive, and
metabolic disorders.
All the tests are performed under standard laboratory procedures, maintaining
proper quality control for accurate results. The Biochemistry Research
Laboratory thus plays a vital role in diagnosis, treatment, and research in the
field of medical biochemistry.
TESTING PARAMETERS IN RESEARCH BIOCHEMISTRY LAB
TEST NAME FUNCTION NORMAL RANGE
TSH ( Thyroid Stimulating Secreted by the pituitary
Hormone ) gland to regulate thyroid 04-4.0 mIU/L
function.

T4 (Thyroxine) Main thyroid hormone 4.5-12.0 µg/dL.

controlling metabolism.
T3 (Triiodothyronine) Active thyroid hormone 80-200 ng/dL.
influencing body
metabolism.

FT4 (Free Thyroxine): The unbound, biologically 0.8-1.8 ng/dL.

active form of T4.

FT3 (Free Triiodothyronine) The unbound active form of 2.3-4.2 pg/mL.

T3.

PRL (Prolactin) Hormone from the pituitary Males : 2-18 ng/mL,

that stimulates milk Females : 2-29 ng/mL.
production and affects
reproductive function.
FSH (Follicle Stimulating Regulates development of males 1.5-124 mIU/mL,
Hormone) ovarian follicles and sperm females 3-20 mIU/mL
formation. depending on menstrual
phase.

LH (Luteinizing Hormone) Controls ovulation and males 1.7-8.6 mIU/mL,

stimulates testosterone females 2-15 mIU/mL
production depending on phase.

Fer (Ferritin) Protein that stores iron and males 30-400 ng/mL,
indicates body iron levels. females 15-150 ng/mL.
PSA (Prostate-Specific Protein made by the
Antigen) prostate; used in prostate < 4.0 ng/mL
screening.
ADVIA CENTAUR CP – MAINTENANCE PROCESS
Perform these tasks at the start or end of each shift.

a) Check Consumables
. Check wash buffer, deionized water, diluent, reaction vessels (RVs), tips, waste
container.
. Refill or replace if low.

b) Clean Probes
. Run Probe Wash program.
. Clean sample and reagent probes with probe cleaning solution.

. Inspect for clots; if present, soak in warm DI water.

c) System Clean

. Run System Rinse or Start-Up Clean Cycle.

d) And reagents & control
Reagent : shake new reagent bottle and added in the correct position ( if light are
blinking )

Control : . Adding control the sequence of . K1 – K2

K2 – K3
K3 – K1
. Go to QC menu
. select run QC
. Choose the control

e) Check Calibrators & Process of the calibrator

Confirm: . Expiry dates
. Sufficient volume
. Refrigerated reagent status (2–8°C)
Different calibrator for different hormone panels ;

• CAL A Used for : FT3

.FT4
 . T3
. T4
Volume required: 500 µL each level (Low & High)

• CAL B Used for: . FSH

. LH
. Prolactin (PRL)
Volume required: 500 µL each level

• CALC Used for: Ferritin (FER)

Volume required: 500 µL each level

• CALQ Used for: PSA

Volume required: 500 µL each level

• CAL3A Used for: TSH-UL (Ultrasensitive TSH)

Volume required: 500 µL each level

• Start Calibration
Press Run Calibration
Machine aspirates calibrators → processes → generates calibration curve.

. Review Calibration Status

After run finishes, check:

✔ "OK / ACCEPTED"

Calibration curve is valid

Test becomes READY

✔ "FAILED / REJECTED"
Possible causes: Expired calibrator , Improper pipetting ,Dirty probe ,Reagent pack expired
,Incorrect calibrator lot
Solution: repeat calibration → check QC afterwards.

. Post-Calibration QC Check
After calibration, always run QC:
Low level QC
Normal level QC
High level QC
If QC passes → Test is ready for patient samples.

Calibration Curve Interpretation

Calibration graph
A good calibration will show: Smooth curve

Tips for Accurate Calibration

✔Use fresh calibrators only

✔ Never shake, only roll gently

✔ Avoid bubbles in cups

✔ Keep probe clean

✔ Use correct lot number

✔ Maintain machine temperature stability

General Sample Handling & Loading:

Specimen Prep: Use serum/plasma follow standard venipuncture, ensure clot formation,
and centrifuge promptly.
Load Tubes: Place barcoded patient tubes into designated sample racks.
Load Racks: Place racks in the sample compartment on the machine.
Software Interface: Log in, go to the workspace, select the assay/sample, and confirm
details.

Result interpretation: The machine interprets the results, compares them to established
ranges, and generates a report

Shutdown Procedure
1. Stop all tests.
2. Remove reagents if needed (refrigerate).
3. Run Shutdown Clean.
4. Turn off analyzer.
Meaning of Replicate (R) and Retest (RE)
✔ Replicate (R)
When a test is performed again on the same day on the same sample, it is called a
Replicate.
Reason: to check precision, repeatability, same-day reproducibility.

➡ Example:
Today you ran FT4 and repeated it again today → Replicate (R).

✔ Retest (RE)
When a test done today but the report is released next day, then that test is called Retest
(RE).
Also used when: The first result is doubtful
. Instrument error
. QC issue
. Sample stored and retested next day

➡ Example:
Today FT4 done → stored → repeated tomorrow before reporting → Retest (RE)
In ADVIA, result flags show:
R = Replicate (same day repeat)
RE = Retest (next day repeat or repeat after storing sample)

Replicate – Retest Calculation (Accuracy Formula)

Step 1 Find the difference
Difference = Higher value - Lower value
Step 2 Divide by the lower value
Index Difference Lower value
Step 3 Convert to percentage
Percentage Error = Index × 100
PASS CRITERIA
If Percentage Error ≤ 10% Result is VALID
If Percentage Error > 10% Result is INVALID
 : CENTRAL LABORATORY :
Introduction
I completed my training in the Central Laboratory, where I gained practical experience in
two major diagnostic departments: Hematology & Biochemistry
During this training period, I learned various laboratory procedures, sample processing
techniques, instrument handling, and quality control measures. This report summarizes
the major tests, principles, and skills I acquired.
1. HEMATOLOGY DEPARTMENT
During my training in the Hematology Department, I learned to analyze blood
components, perform routine hematology tests, and operate automated analyzers.

PT , APTT & INR Value measurement

PT ( Prothrombin Time )
Function / Purpose:
Measures the time required for blood plasma to clot.
Evaluates Extrinsic pathway and Common pathway of coagulation (Factors I, II, V, VII, X).
Important for:
Liver disease monitoring
Vitamin K deficiency
Anticoagulant therapy
Bleeding disorders
Procedure : PT Procedure done on ERBA ECL 412 machine

APTT ( Activated Partial Thromboplastin Time )

Function / Purpose: Measures time taken for intrinsic & common pathway clotting (Factors
XII, XI, IX, VIII, X, V, II, I).
Used to diagnosis Hemophilia A & B
DIC
Coagulation factor deficiency
Monitor Heparin therapy
Procedure : APTT Procedure done on ERBA ECL 412 Machine
INR ( International Normalized Ratio )
What is INR?
INR is a standardized value used to interpret the PT (Prothrombin Time) test results.
It helps compare PT results between different labs, even if the reagents or machines are
different.

✔ Why INR is Needed?

Different labs use different thromboplastin reagents → PT values vary.

To avoid confusion, PT results are converted into INR, which remains uniform worldwide.

Test name Normal range Purpose Clinical use

PT ( Prothrombin 11 – 14 sec Measure speed of Liver disease ,
Time ) clot formation using vitamin K
tissue factor deficiency.
APTT ( Activated 28 – 40 sec Measures clotting Hemophilia , DIC ,
Partial time using activator Coagulation factor
Thromboplastin and phospholipid deficiency ,
Time )
INR ( International 0.8 – 1.2 Standardizes PT Adjusting warfarin
Normalized ratio ) result across dose , bleeding /
different labs clotting risk
assessment

ERBA ECL 412 MACHINE PRINCIPLE

The ERBA ECL 412 works on the Photo-Optical Clot Detection Principle.

✔ Photo-Optical / Turbidimetric Clot Detection

The machine passes LED light through the reaction cuvette containing plasma + reagent.
During clot formation, fibrin strands increase the turbidity (cloudiness) of the sample.
This reduces the amount of light passing through.
The optical sensor detects the change in light intensity.
When the turbidity reaches a defined point, the machine stops the timer.
The time taken = Clotting Time (PT / APTT )
PT PROCEDURE & PRINCIPLE ON ERBA ECL 412
Principle of PT Test:
Plasma is mixed with Thromboplastin reagent (contains tissue factor + calcium).
→ This activates the extrinsic pathway, and the machine measures clotting time in seconds
based on light detection or mechanical clot formation.

PROCEDURE :
1. Collect blood in 3.2% sodium citrate tube (blue cap).
2. Centrifuge → separate platelet-poor plasma.
3. Load plasma sample into ERBA ECL 412 sample rack.
4. Load PT reagent (Thromboplastin) into reagent compartment.
5. Machine aspirates:
A fixed volume of plasma
Pre-warmed PT reagent
6. Reagent is added → clotting starts.
7. Machine detects clot by optical LED sensor.
8. Result displayed in: Seconds
. INR & Prothrombin ratio
APTT PROCEDURE & PRINCIPLE ON ERBA ECL 412
PRINCIPLE :
Plasma is incubated with APTT reagent (contains phospholipid + activator).
→ Then calcium chloride is added to start clot formation.
→ Machine measures the time required to form a clot using optical detection.

🔬 APTT Procedure
1. Collect sample in sodium citrate tube (blue top).
2. Centrifuge → get plasma.
3. Load plasma sample into machine.
4. Load: APTT reagent
. Calcium chloride (CaCl₂) reagent
5. Machine performs: . Pre-warming of plasma
. Adds APTT reagent
.Incubation period
. Adds CaCl₂
6. Clot formation occurs.
7. Machine detects clot optically.
8. APTT result shown in seconds.
Control quality :
Control ( Pro – N & Pro – P ) are run to ensure reagents are working properly .Results are
within acceptable control range .
 : LEISHMAN STAINING :
Used for making Peripheral Blood Smear (PBS) to study:
✔ RBC morphology

✔ WBC differential count

✔ Platelet estimation

✔ Malarial parasite detection

🔷 PRINCIPLE OF LEISHMAN STAINING

Leishman stain is a Romanowsky stain containing:
. Methylene Blue (basic dye) – stains acidic components like:
→ Nucleus (DNA, RNA) = blue / purple
. Eosin (acidic dye) – stains basic components like:
→ Hemoglobin & cytoplasm = pink / red
. The stain contains methyl alcohol, which:
1. Fixes the blood smear
2. Helps dyes penetrate cells
. When the stain is added, methylene blue and eosin react with different cell components
based on their acidic or basic nature, producing characteristic colors.
PROCEDURE OF LEISHMAN STAINING
1)Preparation of Blood Smear
. Place a small drop of blood on a clean slide.
. Use a spreader slide at 30–45° angle.
. Make a thin smear with feathered edge.
. Air dry completely.
2. Fixation & Staining
. Cover the smear with Leishman stain
. Ensure entire smear is covered
. Stain contains methanol → automatic fixation
. Time: 1–2 minutes
3. Add double amount of buffer water
.This dilutes stain and allows proper staining
.Mix gently by blowing or shaking the slide gently
. Time: 8–10 minutes
4. Washing
Gently wash the slide with distilled water
Allow stain to flow off without touching smear
Do NOT over-wash (may remove cells)
5 . Drying
Let the slide air dry in vertical position
Do NOT wipe
6. Microscopy
Examine under oil immersion (100x)
WBC differential count
Parasites (malaria)

7. PRECURTION
Slide must be dry before staining ,
Do not use tap water (chlorine affects staining)
Over-staining → too blue
Under-staining → too pale
pH of buffer water should be 6.8 – 7.2
8 ) Leishman Staining – Results / Observations
. White Blood Cells (WBCs)
Each type has a characteristic appearance:
. Neutrophils
. Lymphocytes
. Monocytes
.Eosinophils ;
.Basophils
2. Platelets
Appear as small purple dots

NORMAL RANGE ;
TEST PARAMETER NORMAL RANGE
WBC (Total Leukocyte Count) 4,000 – 11,000 /µL
Neutrophils 40–75%
Lymphocytes 20–40%
Monocytes 2–10%
Eosinophils 1–6%
Basophils 0–1%
Platelet Count 1.5 – 4.5 lakh/µL
(150,000 – 450,000/µL)

CBC ( COMPLETE BLOOD COUNT ) TEST

CBC (Complete Blood Count) is a blood test that measures the number and
quality of blood cells in your body.
It is one of the most common and important tests done in labs. A CBC test
about Red Blood Cells (RBC),White Blood Cells (WBC) ,Platelets (PLT)
,Hemoglobin (Hb) ,Hematocrit (PCV) ,MCV, MCH, MCHC
CBC Test done by SYSMEX 500 Machine .
Procedure of CBC test in SYSMEX 500 Machine
1. Mix EDTA blood sample by gentle inversion.
2. Open the sample rotor door.
3. Place sample tube in the holder.
4. Select mode:
CBC + DIFF
CBC only
5. Press START / RUN.
6. The analyzer performs: Aspiration
. Dilution
. Lysis
. Counting
7. Wait for the system to complete analysis (≈ 60–90 second )
NORMAL RANGE
TEST NAME NORMAL
RANGE
RBC Count Male: 4.5–5.5
million/µL
Female: 4.0–
5.0
million/µL
Hemoglobin (Hb) Male: 13–17
g/dL<>Femal
e: 12–15 g/dL
PCV / HCT 36–45%
MCV 80–100 fL

MCH 27–32 pg

MCHC 32–36 g/dL

RETICULOCYTE ( RETIC COUNT )

Principle
Reticulocytes are immature red blood cells that contain residual RNA that gets
stained with supravital dyes , forming a visible reticulum , allowing
differentiation from mature red blood cells .

Procedure
Reagents
New Methylene Blue stain (1% solution)
EDTA blood sample (fresh)
Microscope, slides, immersion oil
Steps
A. Mixing and Staining
1. Take 1 drop of EDTA blood and 1 drop of NMB stain (ratio 1:1).
2. Mix gently and incubate for 10–15 minutes at room temperature.
B. Smear preparation
3. After incubation, prepare two thin blood films.
4. Air dry the slides.
C. Microscopic examination
5. Examine under oil immersion (100x).
6. Count 1,000 RBCs and the number of stained reticulocytes.
Reticulocyte Count Calculation
Retic % = (Number of reticulocytes / Number of RBCs counted) × 100
Example:
If 25 reticulocytes are seen in 1000 RBCs:
Retic % = (25/1000) × 100 = 2.5%
Normal Ranges
Adults - 0.5 – 2.5%
Children - 0.5 – 3%
Newborns - 3 – 7%

G6PD ( Glucose-6-phosphate Dehydrogenase ) TEST

PRINCIPLE :
The G6PD test is done to check G6PD enzyme deficiency in red blood cells.
G6PD in RBC converts Glucose-6-Phosphate → 6-Phosphogluconate producing
NADPH.
NADPH reduces blue dye DCPIP into colorless form.
Key Point:
Normal G6PD: Fast decolorization → blue color disappears.
Deficient G6PD: Slow or NO decolorization → remains blue
PROCEDURE:
. Use EDTA Whole blood
. Estimate Hemoglobin content gm/di of whole blood. If the Hemoglobin content is
significantly less than content less than 15 gm/dl, adjust the Hemoglobin content to
15gm/dl by proportionately increasing the aliquat of whole blood during preparation of red
cell hemolysate
. Given below is a table showing quantity of blood required for 1 ml lysing reagent
corresponding to the Hemoglobin concentration Hb gm/dl

Hemoglobin Concentration ( gm/ dl) Quantity of blood to be taken( ml )

7.0 – 9.5 0.04

9.6 – 11.5 0.03
11.6 – 13.5 0.025
13.6 – 15.0 0.02

G&PD-3 (Pre-cooled lysing reagent ) 1.0 ml

Fresh whole blood 0.02 ml

Mix well and keep it in the refrigerator (2-4°C) for 10-15 minutes and use as given below:
1. Transfer completely the red cell hemolysate to the freshly prepared working reagent and
shake well
2.Immediately overlay 1.0 ml. of G&PD-4 Inert Oil
3.Seal the vial tightly, using the plug and cap to make it air fight, incubate at 37C.
4. Observe the change of initial blue to brownish colour.
OBSERVATIONS
Observe the reaction mixture at 30 minutes for decolourisation. the decolourisation is
incomplete, observe for every 5 minutes for shorter intervals thereafter until the
decolourisation is complete. If the decolourisation takes longer time than 10 minutes,
increase the interval time between observations and follow up for 4-8 hours or more.
 : Forward Grouping on Slide:
Purpose:
Forward grouping (cell grouping) is performed to detect A, B, and D (Rh) antigens present on
the surface of a patient’s red blood cells. This test helps determine the patient’s ABO blood
group and Rh (D) type.

Principle
Specific antisera (Anti-A, Anti-B, and Anti-D) react with their corresponding antigens
present on red cells.
Agglutination indicates the presence of the corresponding antigen.
No agglutination indicates the absence of that antigen.

Procedure
1. A clean glass slide was taken and divided into three circles.
2. Three drops of patient blood were placed—one in each circle.
3. The following reagents were added:
. Anti-A on the first drop
. Anti-B on the second drop
. Anti-D on the third drop
4. Each drop was mixed thoroughly with a separate applicator stick.
5. The slide was gently rocked for 1–2 minutes.
6. The reactions were observed for agglutination.

Result / interpretation ;
ANTI - A ANTI - B ANTI -D BLOOD GROUP
+ - + A POSITIVE
+ - - A NEGATIVE
- + + B POSITIVE
- + - B NEGATIVE
+ + + AB POSITIVE
+ + - AB NEGATIVE
- - + O POSITIVE
- - - O NEGATIVE
 :REVERSE BLOOD GROUPING ON TUBE METHOD:
Purpose / Principle
Reverse grouping detects the presence of naturally occurring antibodies (Anti-A and Anti-B)
in the patient’s serum/plasma.
Group A persons have anti-B in their serum.
Group B persons have anti-A.
Group O have both anti-A and anti-B.
Group AB have no antibodies.
The patient’s serum is tested against known reagent red cells:
A cells
B cells
Agglutination pattern helps confirm the forward grouping.

Procedure
At first for Preparing A Cells & B cell
1. Take 2–3 drops of group A and B blood in a test tube.
2. Add 10 mL normal saline.
3. Mix gently and centrifuge at 1500 rpm for 1 minute.
4. Discard the supernatant.
5. Repeat the washing 3 times to remove plasma and antibodies.
6. After final wash, add normal saline to make a 2–5% RBC suspension.
7. Label the tube as A Cells & B Cell (2–5%).
Steps for grouping
. Add 1 drop of patient serum into each tube.
. Add 1 drop of prepared A cells to the first tube.
. Add 1 drop of prepared B cells to the second tube.
. Mix gently. And Incubate at room temperature for 5 minutes.
. Centrifuge at 1000–1500 rpm for 1 minute.
. Gently shake to resuspend the RBC button.
. Check for agglutination and record the result
RESULT AND INTERPRETATION ;

A CELL B CELL ANTIBODY PRESENT BLOOD GROUP

Agglutination No agglutination ANTI - A B
No Agglutination Agglutination ANTI -B A
Agglutination Agglutination ANTI – A + ANTI -B O
No Agglutination No Agglutination NONE AB

:COOMBS TEST :
COOMBS TEST (ANTIGLOBULIN TEST) , There are two types:
1. Direct Coombs Test (Direct Antiglobulin Test – DAT)
2. Indirect Coombs Test (Indirect Antiglobulin Test – IAT)

✅ 1. DIRECT COOMBS TEST (DAT)

Purpose:
To detect antibodies or complement already attached to the patient’s RBC surface.
Used in: Hemolytic anemia
Hemolytic disease of newborn (HDN)
Transfusion reactions
Principle:
Patient’s red cells may be coated with antibodies (IgG) or complement. When Coombs
reagent (Anti-human globulin) is added, it binds to these antibodies → causing
agglutination, which is a positive DAT.
Procedure :
. Take 200 µl of EDTA blood in a test tube add 0.85 % NaCl solution .
. Mix well and wash it three times by centrifuge ( 2000 rpm for the 3 times )
. Finally completely discard the supernatant
. prepare 5% red cell suspension in another test tube by taking 50 µl of washed cells and 950
normal saline solution .
. Mix well and wash it three times by centrifuge .
. completely discard the supernatant
.Add one drop of 5 % washed red cell and two drops of coombs reagent in another test tube
.Mix well and incubate for 15min at 37 degree, after incubator, centrifuge it 1000 rpm for 2
min.
.Examine macroscopically and microscopically
RESULT AND INTERPRETATION
Positive: Agglutination seen → antibodies/complement present on RBC surface.
Negative: No agglutination → no coating of RBCs.

INDIRECT COOMBS TEST (IAT)

Purpose:
Detects free antibodies in the patient’s serum (not attached to RBC).
Used in: Cross-matching before transfusion
Antibody screening
Antenatal testing for Rh incompatibility
Principle:
Patient’s serum is incubated with reagent red cells → antibodies (if present) bind to them.
After washing, Coombs reagent is added.
Agglutination = antibodies present in serum.
Procedure :
.Take patient's clot vial(serum)
.600ul of 'O' positive blood (EDTA) from minimum 3 different blood vials and wash 3 times
.Prepare 5% cell suspension (950ul of NS +50ul of washed cells)
. Take 3 test tubes and label them as T,PC and NC (T-Test serum of 100ul :PC-anti d serum of
2 drops NC-NS of100μ)
.Add 5% cell suspension of pooled 'o' positive (100ul) cells in each test tube
.Incubate at 37 degree celsius for 1 hour
.Add two drops of coomb's serum (anti-human globulin) to each test tube
. Incubate of 37 degree celsius for 15 minutes
.Centrifuge at 1500 rpm for 1 minute
.Resuspend the cells and examine microscopically and macroscopically
Results (IAT):Positive: Agglutination → antibodies present in serum (clinically significant).
Negative: No agglutination → no detectable antibodies.
 BIOCHEMISTRY IN CENTRAL LABORATORY
Introduction :
The Biochemistry Laboratory performs various biochemical, hormonal, liver, kidney, lipid,
electrolyte, cardiac and metabolic tests using an automated . Biochemistry Lab utilizes the
Sys 400 automated analyzer to perform a wide range of tests with high accuracy.
The workflow includes sample processing, reagent handling, analysis, QC, and report
validation.

Common Tests Done in Sys 400 With Normal Ranges

Below is a complete list grouped by test categories:
A. Liver Function Tests (LFT)

Test Name Normal

Range

Total Bilirubin 0.3 – 1.2

mg/dL

Direct Bilirubin 0.0 – 0.3

mg/dL

Indirect Bilirubin 0.2 – 0.9

mg/dL
SGPT/ALT Male: 10–
45 U/L, Female: 7–35 U/L

SGOT/AST Male: 10–

40 U/L, Female: 9–32 U/L

ALP 44 – 147
U/L

Total Protein 6.0 – 8.3

g/dL

Albumin 3.5 – 5.5

g/dL

Globulin 2.5 – 3.5

g/dL

A/G Ratio 1.0 – 2.2

Kidney Function Tests (KFT / RFT)

Test Name Normal Range

Blood Urea 15 – 45 mg/dL

Creatinine Male: 0.7 – 1.3 mg/dL, Female: 0.6 – 1.1 mg/dL

Uric Acid Male: 3.4 – 7.0 mg/dL, Female: 2.4 – 6.0 mg/dL

Sodium (Na+) 135 – 145 mmol/L

Potassium (K+) 3.5 – 5.1 mmol/L

Lipid Profile
Test Name Normal Range

Total Cholesterol < 200 mg/dL

Triglyceride < 150 mg/dL

HDL Male > 40 mg/dL, Female > 50 mg/dL

LDL < 100 mg/Dl

VLDL 5 – 40 mg/Dl
Cardiac Tests
Test Normal Range

CK-MB < 25 U/L

LDH 140 – 280 U/L

Diabetes Profile
Test Normal Range

Fasting Glucose 70 – 110 mg/dL

PP Glucose < 140 mg/dL

Enzymes & Others
Test Normal Range

Amylase 25 – 125 U/L

Lipase 10 – 140 U/L

ACR ( Albumin – creatinine Ratio test )
TEST NAME NORMAL RANGE
ACR VALUE < 30 mg / g

VITAMIN D TEST
PROCEDURE
.Set the temperature to 37 degree before 10 min to prior to starting the test.
.Insert the test device to the test slot of the analyzer. The analyzer read the barcode data
and check the test device is valid .
.Take out a reaction tablet 1 tube from the bottle.
.Collect 175 µl of buffer from the buffer bottle and dispense it into the reaction tablet 1 tube
after add 35µl serum
.Mix thoroughly and incubate the reaction tablet 1 tube at 37 degree for 30 min.
.Take out tablet 2 and mix it wuth incubated reaction tablet 1
.Mix thoroughly and dispense it to the specimen well of the test device .
.Mount the test device ti the lower heat block and incubate it 15 min
.Then insert the incubated test device . The analyzer will immediately display the test result.

Result
Normal : 30 – 100 ng / ml

URINE ROUTINE EXAMINATION

Urine Routine Examination – Dipstick Method
Principle
The dipstick method is a semi-quantitative test where a plastic strip containing chemical
reagent pads is dipped into urine.
Each pad changes color according to the presence and concentration of specific
substances (e.g., glucose, protein, ketones).
The color is then compared with the manufacturer’s color chart to interpret the result.

Procedure
1. Collect fresh midstream urine in a clean container.
2. Mix the sample gently.
3. Remove one dipstick from the bottle (do not touch reagent pads).
4. Dip the strip into urine for 1–2 seconds ensuring all pads are wet.
5. Remove the strip and drag it on the container edge to remove excess urine.
6. Hold the strip horizontally to avoid mixing of reagents.
7. Wait for the recommended time (15 sec–2 min depending on parameter).
8. Compare each pad with the color chart on the dipstick bottle.
9. Record the results for all parameters.

3. Microscopic Examination:
1. Centrifuge urine at 2000–3000 rpm for 5 minutes.
2. Discard supernatant and leave ~0.5 mL sediment.
3. Place a drop of sediment on a clean glass slide. Cover with coverslip.
4. Examine under microscope: o Cells: RBCs, WBCs, epithelial cells
o Casts: Hyaline, granular, RBC, WBC
o Crystals: Uric acid, calcium oxalate, triple phosphate, etc.
o Organisms: Bacteria, yeast, parasites
 ABG Test

INTRODUCTION
The cobas b 221 procedure for ABG (Arterial Blood Gas) involves a fully automated process
where a blood sample (arterial, venous, capillary) is introduced into the analyzer, which
then measures key parameters like pH, PaO2, PaCO2, electrolytes (Na+, K+, Ca2+, Cl-),
hematocrit, glucose, lactate, and hemoglobin derivatives, providing rapid, accurate results
for critical care diagnosis, often using specialized electrodes and spectrophotometry for
different analytes,

PROCEDURE
Step-by-step ABG procedure in Cobas b 221
1. Check machine status
Ensure calibration is complete
QC values within range
2. Prepare the sample
Mix the arterial blood gently (do not shake)
Remove air bubbles from syringe
Keep sample on ice if delay > 5 minutes
3. Load sample into analyzer
Open sample port
Insert the syringe/tube in the sample holder
Machine automatically aspirates the sample
4. Automatic analysis
Analyzer measures pH, pCO₂, pO₂, electrolytes, glucose, lactate
Performs hemoglobin and saturation calculations
5. Result generation
Results appear on screen within seconds
Print or transfer electronically
6. Post-analysis
Remove the syringe
 Dispose safely (biohazard)
Machine performs automatic cleaning

RESULTS PROVIDED
Blood gas: pH, pCO₂, pO₂, HCO₃⁻, BE, TCO₂, sO₂
Electrolytes: Na⁺, K⁺, Cl⁻, Ca²⁺
Metabolites: Glucose, Lactate
Hematology: Hb, O₂Hb, COHb, MetHb, tHb
 STOOLE OCCULT BLOOD EXAMINATION
Occult blood test which is detect hidden blood in the stool as a screening method

.Procedure :
.Special paper or a clean container is used to catch the stool sample without it contacting
the toilet water or urine.
.A small wooden or plastic stick (sometimes with a corkscrew spiral tip) to collect a small
amount of the stool specimen.
: A tube containing a liquid buffer solution into which the collected stool sample is placed
and mixed. The buffer helps suspend and preserve the sample for testing.
.Test Card/Cassette/Strip: This is the actual test mechanism.
. use a card coated with the chemical guaiac, and a developing solution is added later in a
lab to check for a color change.

Result :
If the colour is change indicate the presence of blood in the stool
No colour change indicates the absence of blood
 SAMPLE COLLECTION

INTRODUCTION
In the Outpatient Department (OPD), blood samples are collected based on the doctor's
referral for various laboratory tests. Proper handling, identification, and processing of
samples are essential to ensure accurate and reliable test results.

Role of the OPD Sample Collection Unit

The OPD sample collection area serves as the first point of contact between patients and
laboratory services. The main functions include:
• Receiving test requisition forms from patients.
• Collecting blood samples as per the doctor’s prescription.
• Ensuring proper labeling, handling, and transportation of samples to the laboratory.

. Blood Collection Procedures

Blood samples are collected following standard venipuncture techniques:

1. Patient preparation and verification.
2. Selection of appropriate vein.
3. Use of aseptic precautions.
4. Collection of blood into specific tubes depending on the required test.
5. Gentle mixing of tubes containing anticoagulants.
6. Proper storage and timely dispatch to the laboratory
 Types of Blood Collection Tubes Used
Various tubes are used in OPD sample collection, each with a specific purpose:
1. Sodium Citrate Tube (Light Blue Top)
Additive
:Contains 3.2% or 3.8% Sodium Citrate as an anticoagulant.
Works by binding calcium, which is necessary for clotting.
Purpose / Uses:
• Used mainly for coagulation studies, including:
o Prothrombin Time (PT)
o Activated Partial Thromboplastin Time (APTT)
Special Notes:
• Must be filled exactly up to the mark—incorrect volume affects clotting test results.
• Gently invert 4–5 times after collection to mix.

2. Clot Activator Tube (Red or Gold Top)

Additive:
• Contains no anticoagulant (plain tube) or a clot activator (silica particles).
• Blood clots naturally inside the tube.
Purpose / Uses:
Used for tests requiring serum, including:
• Biochemistry tests
o Liver Function Test (LFT)
o Kidney Function Test (KFT)
o Lipid Profile
., Urea, Creatinine
• Hormone assays
• Serology (e.g., HIV, HBsAg, VDRL)
Special Notes:
• Must allow blood to clot for 20–30 minutes before centrifugation.
 • Provides clear serum for testing.

3. EDTA Tube (Purple Top)

Additive:
• Contains Ethylenediaminetetraacetic acid (EDTA).
• Prevents clotting by chelating (binding) calcium.
Purpose / Uses:
Used mostly for hematology, such as:
• Complete Blood Count (CBC)
• Peripheral Blood Smear
• Hemoglobin estimation
• Packed Cell Volume (PCV)
• Blood grouping
• HbA1c
Special Notes:
• Gently invert tube 8–10 times to avoid clots.
• Excess EDTA can cause cell shrinkage, affecting results.

4. Sodium Fluoride (Grey Top)

Additive:
• Contains Sodium Fluoride: inhibits glycolysis (stops glucose breakdown).
• Contains Potassium Oxalate as anticoagulant.
Purpose / Uses:
Specially used for glucose-related tests, including:
• Fasting Blood Glucose (FBG)
• Postprandial Blood Glucose (PPBG)
• Random Blood Glucose (RBS)
Special Notes:
• Prevents decrease of glucose levels after collection.
 • Ideal for accurate sugar results because cells cannot use glucose.

Sodium fluoride EDTA CLOT sodium citrate

Urine sample collection

1.Sample Collection at OPD:
At the OPD, the patient first shows the bill for verification. According to the test mentioned
in the bill, the patient is instructed on how to properly collect the urine sample.
2.Patient Instructions:
The patient is guided to collect a clean, mid-stream urine sample in a sterile container. The
container is properly closed after collection to avoid contamination.
3.Sample Labeling
After collection, the sample container is labelled with the patient’s name, age, sex, hospital
number, test name, and date/time of collection.
4. Sending to Laboratory:
The labelled sample is then sent to the laboratory for further processing and testing.

Semen sample collection :

Semen Sample – Same Procedure:
For semen samples, the process is similar. The patient shows the bill, receives collection
instructions, provides the sample in a sterile container, and the sample is labelled and sent
to the lab.