MODEL PAPER -1 n 1

MODEL PAPER -1
DIPLOMA IN MEDICAL LAB TECHNICIAN
SECOND YEAR
PAPER - I
Time : 3hrs. Total Marks : 80

SECTION – A

Note: Answer any Ten of the following.

4 × 10 = 40 M
1. Write about the cell structure division and
functions of cell organelles?
2. Write about the gross and histological struc-
ture and function of liver.
3. What is fixation? Explain about varying fixa-
tion types.
4. Write about museum methods of specimen
preservation.
5. Explain about sharpening of microtome
knife?
6. What is embedding? Write about tissue em-
bedding systems?
7. Write about fine needle aspiration.
8. Write about the maintance of the stains?
9. Explain about formation of blood?
10. Write about collection of blood?
11. Write in detail regarding blood banking
12. Write the following.
1. Anticoagulanta
2. Blood cell count.

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ANSWERS
SECTION – A
1. Write about the cell structure division and
functions of cell organelles?
Ans: The cell is the basic structural and functional
and biological unit of all known living or-
ganisms.
Cells are the smallest unit of like that can
replication independently and are often
called the building blocks of life. The study
of cells is called cell biology. The cell was
discovered by Robert Hooke in 1665.
Cells consists of a Protoplasm enclosed
within in membrane, which contains many
biomolecules such as proteins and nucliec
acids.
The interior of the cells is divided into the
nucleus and the cytoplasm. The nucleus is
a spherical and oval shaped structure of the
cell. The cytoplasm is the region outside the
nucleus that contain cell organelles & Cyto-
sol, cytoplasmic solution.

Figure : Structure of cell

a.) Cell membrane (or) Plasma membrane:

It is a protective sheath of cell body. It sepa-
rates the fluid outside the cell called extra
cellular fluid and the fluid inside the cell is
called intracellular fluid.
Plasma membrane consists of
Lipids
(Phospholipidse cholesterol)
Proteins
Carbohydrates
b.) Cytoplasm : It is a fluid present inside of
the cell. It contains a clear liquid portion
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 MODEL PAPER -1 n 3

called cytosol. It constitutes about 55% of movement of granules, vesicles and protein
total cell volume. molecules.
c.) Cell Organelles: These are specialised struc- • Gives structural strength to the cell.
tures within the cell that have characteristic
shape which perform specific functions in • Provides resistance to the cell
cell growth, maintenance and reproduction. Centrosome : ® located near the nucleus
consisting of two components i.e., a pair of
Cytoplasmic organelles are
centrioles and pericentriolar material. The
• Cytoskeleton, two centrioles are cylindrical structures and
• Ribosomes, each composed of nine clusters of 3 micro-
tubules arranged in a circular pattern.
• Endoplasmic reticulum,
Cilia ® These are the nobile projection of
• Golgi apparatus, the cell surface, cilia are numerous, short
• Lysosomes, hair like projections that extends from the
surface of the cell.
• Peroxisome,
Flagella ® These are similar in structure
• Centrosome,
to cilia,but are much longer. It moves on
• Centrioles, secretory vesicles, the entire cell.
• Mitochondria and nucleus. Endoplasmic Reticulum : It consists of tubu-
lar and vesicular structures arranged in the form
1. Cytoskeleton : Cytoskeleton of a cell is a com-
of interconnected network in the cytoplasm.
plex network of structures in various sizes
present throughout the cytoplasm. It determines Endoplasmic reticulum forms link between
shape and gives support to the cell. The cy- nucleus and cell membrane by connecting the
toskeleton consists of 3 major protein compo- cell membrane with nuclear membrane.
nents which are -
This two types : Rough E.R
a) Micro Tubules : These are straight and hol-
Smooth E.R
low tubular structure without limiting mem-
brane arranged in different bundles. The • Rough ER : Rough ER is vesicular in
assembly of microtubules begins in an or- structure. Granular ribosomes are attached
ganelle called the centrosome. to outer surface ® gives a giving the bread-
like appearance. It helps in synthesis of
b) Intermediate Filaments : It forms a net-
proteins in the cell.
work around the nucleus and extend to the
periphery of the cell. Diameter of each fila- • Smooth ER: ® does not have ribosomes.
ment is about 10 nm. It helps to maintain It is formed by many interconnected tu-
the shape of the cell shape. bules. So, it is called tubular ER. It is re-
c) Micro Filaments : These are long and fine sponsible for the synthesis of non-protein
thread like structure with a diameter of 3 to substances such as cholesterol and steroid.
6nm. They are composed of actin and myo- • Golgi Apparatus : Situated near the
sin. nucleus. Each golgi apparatus consists of 5
Functions: to 8 membranous sacs. These sacs are usu-
ally flattened and are called cistemae.
• Responsible for movements of centrioles.
Functions : It helps in processing and de-
• It acts like convey belts which allow the livery of proteins and lipids.
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Figure : Golgi apparatus complex a) Ribosomes ® are granular dot like

structures with a diameter of 15 nm.
Lysosomes: These are membrane-bound These are made up of 35% proteins and
vesicular organelles found throughout the 65% Ribo Nucleic Acid (RNA).
cytoplasm. These are formed by golgi ap-
paratus. These are called suicidal bags. Functions ® help in synthesis of pro-
teins.
Functions : Its main function is phogocy-
tosis, that carries on two ways such as: Nucleus ® It is a membrane-enclosed or-
ganelle found in eukaryoticcells. lt contains ge-
• Heterophagy ® which means digestion of
netic material, large variety of proteins, such as
extra cellular materials engulfed by the cell.
histones, to form chromosomes.
• Autophagy ® which means digestion of
Function ® to maintain the integrity of these
intracellular substances such as worn out cy-
genes and to control the activities of the cell by
toplasmic organelles.
regulating gene expression.
Peroxisomes : These are the membrane lim-
The main structure making up the nucleus are
ited vesicles like the lysosomes. These are
nuclear envelops
formed from ER. These contain some en-
zymes such as catalase, oxidase. Functions : Synthesis of RNA
Functions: Control the cell division
• Degrades toxic substances like hydrogen Control all the activities of cell
peroxide and other metabolic end products Storages of heriditiy information
by means of detoxification.
Chromosomes consists of genes
• Breakdown of excess fatty acids. that contro cellular structures &
Mitochondria: It is a oval-shaped structure direct cellular function.
with a diameter of 0.5 to 1m . It is bilayered 2. Write about the gross and histological
membranous organelle. Outer membrane structure and function of liver.
is smooth and the inner membrane is folded
in the form of shelf-like inward projections Ans : Liver is a large, solid gland sitiuated in the
called cristae. Cristae contain many enzymes right ipper quadrant of the abdominal cavity.
and other protein molecules which are in- The liver is the largest gland in the body. It
volved in respiration and synthesis of ATP. secrets bile performs metabolic functions.
Functions: Shape ® Wedge shaped
• It is called power houses of cell because, it Colour ® Reddish brown
produces the energy required for cellular
Weight ® Males ® 1600gr
functions.
females ® 1300 gr
• Synthesis of ATP
Location ® Liver occupies the greater part
of the right hypochondril and it is a part of
epigastrium and extends into the left
hypochardium.

Figure : Golgi apparatus complex Relations :

It consists of Superiorly ® Diaphrams anterior abdomi-

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 MODEL PAPER -1 n 5

nal wall. It has caudate lobe ® situated on posterior

surface it is bound by inferior venacova and
Inferiorly ® Stomach, bile ducts duodenum ligamentum venorm.
adrenalgland & kidney.
• Quardate lobe : Seen on ingerior sur-
Posteriorly ® Esophagus, inferior venacova face rectangular shape. It is formed by
aorta gall bladder and vertebral column inferior or border ,portahelpats and fossa
Laterally ® Lowe ribs is diaphgrams for gall bladder in left by fissure. for ligam-
entumteres,
Figure : Location & parts of liver
b.) Left lobes : It is smaller than the right lobe it
Surface of Liver : looks flattened and form the above down-
wards.
• Anterior surface ® It is triangular and
slightly convex Histology :
• Posterior surface ® It lies between poste- • Liver is covered by Glisson’s capsule.
rior superiors posterior inferior boarders, • Sheets of connective tissue divided the liver
which are not well defined. into thousands of small units, it is called lob-
• Super surface ® It is quardilateral and in ules. The lobules is hexogonal in shape, with
the midline, it should a concavity. portal triads at the verticles and central vein
in the middle.
• Inferior surface ® It shows following fea-
tures from left to right. The actions of liver is defined as the liver
parenchyma around a pretermination
• Fissue ligamentumteres branch of hepatic arteriole between two
• Fossa for gallbladder adjacent central veins. It is the function unit
of lever.
• Caudate and papillary process of caudate
lobe. Impression for colon & renal etc., The parenchymal cells of the liver are hepa-
tocytes they make contract with blood in
Boarders of liver sinusolids.
• Inferior boarder ® is sharp arteriorly where Bile orginates as secretions from the basal
it separates the anterior surfaces from the surface of hepatocytes which collect in chan-
inferior surface. nels called canaliculi.
• Posterior superior boarder ® It is demov - These secretions of low towards the periph-
cated by superior layer of coronary ligament, ery of lobular and into bile ductablar and
upper ends of groove for inferior venacova interlobular bile ducts ultimately collecting
and left triangular ligament. in the hepatic duct outside the liver.
• Posteroir inferior boarder : It is indicated by • Arterial Supply ® liver recevies 80% of
the layer of coronary ligament and groove its blood supply through the hepatic ar-
for the inferior venacova and also separates tery & 80% through the portal vein. Be-
the inferior and posterior surfaces lobes of fore entering the liver both hepatic and
liver. portal vein divide into right and left
a.) Right lobes : It is much larger than the left branches.
lobe and forms 5% 6 of the liver. It contrib- • Venous driange ® hepatic sinusoids
utes to all the 5 surfaces of liver and present drain into interolobular veins, which joins
the caudate and quadrants lobes. to form sublobular veins. These inform
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unit to form hepatic veins which drain • Anhydrous disodium phosphate

directly into inferior venacava. 0.65g.
• Lympathic driange ® Superficial lym- • Distilled water 100ml
phatic of the liver run on the surface of c.) Formal calcium : (Lillie 1965)
the organ beneath the peritenium amd
ferminate in canal, hepatic paracardiac • Formalin 10ml
a coleliac lymph nodes. • Calcium acetate 2.0g
• Nerve supply ® Liver receives nerve • Water 100ml
supply from hepatic flexus, which con-
d.) Buffered formal sucrose (Holts &
tains both symphathetic & parasympa-
Hicks 1961)
thetic (or) vagal fibers.
• Formalin 10ml
3. What is fixation? Explain about varying
fixation types. • Sucrose 7.5g
Ans : Fixation : When a tissue is removed from • M/15, phosphate buffer 10ml
the body it begins to decompose. To pre- (pH 7.4)
serve the natural state the tissue immedi-
Features: Excellent fixative for preservation
ately to be placed into a solution is called
of fine structure phopshate and
fixative process is called fixation.
some enzymes.
Types : Fixatives can be diviedc into the fol-
For cytochemistry & electron mi-
lowing three main groups
croscopic studies
1. Microanatomical fixatives
Should be used cold (4 oC) on
2. Cytological fixatives fresh tissue.
3. Histochemical fixatives e.) Heinden hains susa
1. Microanatomical fixatives: These are • Mercuric chloride 4.5g
mainly used for te routine use. These are
• Sodium chloride 0.5g
used to preserve the anatomy of the tis-
sues, with the correction relationship of • Trichloroacetic acid 2.0g
tissue layers and large aggregates of cells • Aceticacid 4.0ml
following are the examples of micro ana-
tomical fixatives. • Distilled water 100ml
a.) 10% (V/V) formalin in 0.9% sodium Specific features :
chloride (normal saline). It is a rou- • Excellent fixative for routine biopsy.
tine fixatives choice for many years
now been supplemented by buffered • Allow brillan staining wiht good cytologi-
formalin (or) by formal calcium ac- cal detail.
etate. • Gives rapid and even penetration with
b.) Buffered formalin minimum shrinkage

• Formalin 10ml • Tissues should be trated iodine to re-

move mercury pigment.
• Acid sodium phosphate monohy-
drate 0.4g. f.) Zenker’s fluid
• Mercuric chloride - 5g
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 MODEL PAPER -1 n 7

• Potassium dichromate 2.5g a. Nuclear fixatives

• Sodium sulfate 1.0g b. Cytoplasmic fixatives
• Distilled water 100ml a. Nuclear fixatives
• Add immediately before use gla- • Carnoy’s fluid
cial acetic acid 5ml • Chloroform 30ml
Specific features : • Absolute alcohol 60ml
• Good routine fixatives • Galcial acetic acid 10ml
• Gives fairly rapid and even penetration. - Specific features ® It penerates very
g.) Zenker formal (Helly’s fluid) rapidly and gives excellent nuclear fixa-
tion.
• Mercuric chloride - 5g
- Nissal substances and glycogen ae pre-
• Potassium dichromate 2.5mg
served.
• Sodium sulfate 1.0g
- Good fixatives for carbohydrates
• Distilled water 100ml
- It causes considerable shrinkage
• Add formalin immediately before
- It destroys dissolves most cytoplasmic el-
use 5ml
ements.
Specific features :
• Clarke’s fluid absolute alcohol 75ml
• It is an excellent microanalytical fixative
Galcial acetic acid
• It an an excellent fixative for bone mar-
Specific features :
row spleen & blood containing organs.
• This fixatives penetrates rapdily, gives
• As with zenker’s fluid it is necessary to
good nuclear fixations effects preserva-
remove excess dichromates and mercu-
tion of cytoplasmic elements.
ric pigment.
• It is an excellent fixative for smear (or)
h.)Bouin’s fluid
coverships preparation of cell for gen-
• Saturates aqueous picric acid eral fixatives chromosomal analysis.
75ml
• New corner’s fluid
• Formalin 25ml
Isopropanal 60ml
• Glacial acetic acid 5ml
Propionic acid 40ml
• This fixative penetrates everluso
Petroleum ether 10ml
rapidly and causes little shrinkages.
It can be demonstrate glcogen. Acetane dioxane 10ml
• Tissue fixed in it gives brilliant Specific features :
staining by the trichrome methods. • This fixatives penetrates rapidly and pre-
II. Cytological fixatives : These are used when serves the chromatin better then carnoy’s
the preservation of intracellular (or) inclu- fluid. It is food fixature for the presenta-
sions is very important. These fixatives can tion of mucopoly saccharids.
be subdivided into. b. Cytoplasmic fixatives

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• Champy’s fluid. • Organ demonstrations play an important
3g/dl potassium dichromate 7ml role in teaching.

1% (v/v) chromatin acid 7ml • Autopsy laboratories are inevitable to receive

requests to demonstrate gross anatomy or
2g/dl osmium tetroxide 7ml pathology to students in the allied health
Specific features : sciences and also in the primary and sec-
• This fixative cannot be stored hence, it ondary schools.
should be prepared fresh before use. It • Surgical pathology museums are a priceless
penetrates poorly and unevenly. components of any pathology department.
It preserves mitochondria fat, yolk and • The mounting of pathological specimens, in
lipids tissue must be washed overnight addition to their teaching value, play a part
after fixation. in recoding the history of madicine.
III. Histochemical fixatives : • The specimens are mounted in glass on
It is used for histochemical studies. Freeze acrylic jar.
drying tecnique is ideal for this purpose. • Before placing in the jar, they are pinned
Functions: Preservation of the tissue on as acrylic sheet. The specimens are
consistuents . treated with different solutions, which pres-
sures the original shape and the related spe-
Preservation of morphological relationship
cific colours.
Preservation of specific tissue constitutents
& No inteference with the reagents to be Preparation of specimen:
used in the process of visualization bufered 1. Along bladed, sharp knife should be used
formalin is the most common histological (the usual brain knife or a butcher’s knife
fixative. with a 14 inch blade).
• Buffered formalin is the most common 2. Cut surfaces should be even and smooth
histological fixative. The flexatives used (this can be achived by using a continuous
in histopathology laboratory. stroke).
Formaldehyde ® used by hearing at 3. Tissue for histological examination should
80oC for -4hrs. be taken from the back of the specimen or
Glutaraldehyde ® 50% (v/v) aqeous should be removed from the front with a
solution can be 80oC for 2min 4hrs ® at scalpel.
60oC for 7hrs. 4. Specimen should be put into a fixative im-
A crolein (or) chromal chloride ® used in mediately
liquid form at 37oC for 1-2 hrs. 5. As primary fixative 10% (v/v) formal aline
4. Write about museum methods of specimen can be used and later on the specimen is
preservation. transferred to a special fixative.

Ans : Surgical pathology museum is a depository The solutions required for this technique are
of anatomy specimens, their description, as follows:
supporting investigational and clinical details. Formalin : 400 ml.
• Museum represents informaltion of pioneer Potassium nitrate : 30 g.
work of eminent scientists and history of
medicine. Potassium acetate : 60 g.

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 MODEL PAPER -1 n 9

Tap water (filtered) : 2,000 ml. • Honing and

Kaiserling fluid II : 80% (v/v) ethyl alcohol • Stropping.
is used to restore colour in an emergency. Requirements:
Pulvertaft- Kaiserlingimounting fluid III is • A hone (rectangular block of stone cpecially
used as mounting medium for the specimen. made for sharpening of knife).
It is
• A properly fitted back of the knife.
Prepared as follows:
• Microscope to observe the edge of the knife.
• Glycerin : 300 ml.
• Knife.
• Sodium acetate : 100 g.
• A strop (flexible or grid).
• Formalin : 5 ml.
Procedure (Honing):
• Tap water (filtered) : 1000 ml.
• Clean the knife with Xylene soaked cloth.
0.4 g/dl sodium hydrosulfite is added im-
mediately before sealing the jar. • Put on the proper honing back.
Storage of specimen: • Keep a damp cloth under the hone and po-
sition the hone on suitable bench.
1. The specimen, together with a duplicate la-
bel, is wrapped in gauge or muslin and the • Lubricate the surface of hone with coconut
label is attached with piece of linen thread. oil.
2. The specimens are stored in a large rectan- • Place the knife at one end and push it di-
gular earth enware tanks (34 × 22 × 1inches) agonally forward with the leading cutting
containing kaiserling’s fixing solution 1 for a edges in the front.
period up to 6 months. • Just before the edge reaches the end of the
3. It is treated with 80% (v/v) alcohol. stone the knife is turned, over on its back
without lifting it. It is steadily pulled back with
4. The specimen is placed afterwards in the cutting edge leading again along the
pulvertaft Kaiserling mounting fluid (with out hone towards the operator.
sodium hydrosulfite).
Stropping technique
5. Explain about sharpening of microtome
knife?
Ans : Sharpening of the Micro tome Kife:
Importance: The edge of the microtome
kinfe performs the function of cutting and it
should be sharp in order to get good sections
of the tissues. Honing technique
Introduction : Sharpening of the knife can
be carried out by:
• Mechanically or by
• Automated knife charpners.
The process of sharpening consists of:

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Observe the edge of the knife under micro- mould containing paraffin wax. Cut surface
scope to check the progress of the honing. of tissue resets flat on to base of the mould.
Precautions :The pressure on the knife Leuckhart Mould Method: ‘L' shape embed-
should be just sufficient to maintain its edge ding mould is made up of two ‘L’ shaped
in contact with hone surface. pieces of heavy metallic brass. These are
placed on glass plate.
B. Procedure (Stropping):
Glycerin may be applied to the glass plate
Importance: Stropping is done in order to
and ‘L' pieces. Molten wax is poured in the
remove the wire edge caused by the fric-
mould. Tissue is oriented in the mould and
tion of the steel upon the stone during hon-
label is placed on it. Molten wax is allowed
ing. The strop is usually made up of leather.
to cool. On cooling, moulds are seperated.
Procedure: Wax is cut into different blocks seperating
tissues. Wooden or metal holder is attached
• Take a clean and dry knife.
to the block, which is then ready for cut-
• Place a rigid strop on the bench. ting. This is considered as traditional block-
• Place the knife at one end of the strop and ing method 'L’ moulds are used in the labo-
push it diagonally forward carefully so that, ratories with less work load.
entire edge receives the polish. Tissue Embedding Systems: Tissue em-
• Maintain the pressure on the knife just suffi- bedding systems allow easy and simple em-
cient to keep the edge in contact with the bedding of tissue. These systems are com-
surface of the strop and establish a rhyth- prised of the following components:
mic, steady motion. 1. Heated chambers for storage of moulds,
• When it reaches the end of the strop, turn it forceps, and cassette bath.
on its back and pull back towards you (to- 2. Paraffin wax reservoir (3-5 litre capacity).
wards the operator).
3. Wax dispenser.
• Observe under microscope for a sharp and
4. Cold plate adjusted to -5° to-5°temperature.
even surface.
50 to 60 blocks can be placed on it for cool-
6. What is embedding? Write about tissue ing.
embedding systems?
5. Drain tray for excess wax
Ans : Embedding : means casting or blocking. In
this process, the infiltrated and impregnated
tissue is placed in warm liquid paraffin, which
forms a firm block after cooling.

Fig : Tissue embedding System

Embedding is done by filling a metal base

Fig : Paraffin embedding requirements mould of suitable size with molten paraffin .
Specimen is oriented in such a way that it
Embedding enables the tissue to be cut on
will cut in right plane. A plastic is put over
a microtome. Tissues are oriented in base
the mould and it is allowed to solidify. After
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 MODEL PAPER -1 n 11

cooling the metal mound is peeled away and • Infections or inflammatory condition can also
plastic remains attached to paraffin block. be dianosed by FNAC.
The paraffin blocks can be directly clamped • It is necessary to indicate, whenever palpable
to the microtome. mass or lesion is visualised within any or-
Note: gan.
1. Tissue tek moulds are used as base moulds. • FNAC of deep- seated lesions are performed
They are made up of stainless steel and are under radiological guidance using a similar
available in the following different sizes: technique that is used for superficial masses.
7×7 mm, 15×15 mm, • FNAC can be performed in out patient de-
partment (OPD), clinics or in specific room
24×24 mm, 37×24 mm
in nursing homes.
2. Following two components are used in em-
• The sample is received in the form of smears.
bedding:
• Following are the requirements for FNACs
a. Plastic embedding ring : The tissue is
placed in the base moulds and plastic ring is 1. Examination couch with following facili-
placed on the top and wax is poured in the ties:
ring and cooled rapidly. Plastic ring func- • Pillow.
tions as block holder.
• Disposable bed sheet.
b. Embedding cassettes (cassette holder
method): This is a modification of plastic • Blanket.
embedding ring. • Screen for privacy.
Embedding cassettes hold tissue for 2. Solution and equipments:
processiong. Base of the cassette has one
end with slope on which identification num- • Fixative : 80% (v/v) isopropanol.
ber is viewed and is used as block holder. • Formal alcohol.
This is placed on metal base after embed-
ding of tissue. • 10 ml syringes.

Orientation of Tissue: • Needle size 22 g or 23 g.

1. Select appropriate size of base mould for • Sterile gauge and plaster.
embedding. • Gloves.
2. Place tissue in the center of the mouls, which • Glass slides with frosted ends.
will allow paraffin support for the tissue while
• Consent form.
sectioning. At least 1 mm of paraffin should
be present on all sides beyond the tissue. • Staining solution
3. Bone marrow, to cut needle biopsies should Procedure:
be placed horizontally.
• Palpitation of suspected lump: The lump
4. Tabular structures like fallopian tubes, ap- is localised and checked by palpating be-
pendix are embedded transversely tween the tips of the index and middle fin-
gers. The selected area is cleaned with an
7. Write about fine needle aspiration.
alcohol swab.
Ans : Fine needle aspirates are usually obtained
• Aspiration: The needle is inserted through
to diagnose or rule malignancy.

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skin rapidly and then carefully into the lump; 4. Gram stain: For micro organisms.
plunger is pulled back and needle is moved 5. Alcian blue stain: To demonstrate acid
in straight line with back and forth motion mucins.
and plunger is released before with draw-
ing needle. 6. Alcian blue PAS stain: To distinguish acid
and neutral mucins.
• After with drawing the needle a gauze piece
is applied over the puncture site with pres- 7. PAS stain: For fungus, differentiate between
sure. adenocarcinoma and mesothelioma using
diastase digestioh technique.
• After removing the needle the collected ma-
terial is placed on the glass slides. 8. Oil Red ‘O’: To identify fats in lipid
pneumonia.
• Smear preparation: Smears are prepared
by using glass slides. • Filter stains and reagents daily.
• Smears are immediately stained with tolui- • Non- Gyn specimens have high potential
dine blue. for cross contamination during staining
procedures.
8. Write about the maintance of the stains?
• Solutions and stains are changed from
Ans : Following precautions are taken while using time to time depending on the volume
the stains: of work.
• Stains used must be certified by biological • Stains are stored in the dark bottle when
stain commission. not in use.
* Dye content on the certified label of dye must • Staining dishes should be kept covered.
be considered in stain preparation.
• OG- EA solution loses strength rapidly
* Contamination control: Gynec and Non- and should be replaced as soon as cells
gynec slides are stained seperately. Cells are appear grey dull or with out crisp con-
detached from the slides during staining. trast.
These cells can cause cross contamination
which results in false diagnosis. • Alcohol used during rehydrating and de-
hydrating process are changed on a ro-
Special stains used for smears: tation basis and if needed also filtered
1. ZNstain: For acid bacilli. prior to staining procedure.
2. GMS stain: For pneumocystis carinii, fungus. • Daily microscopic checks are docu-
mented for the quality control of stains.
3. Mucicarmine stain: For cryptococci and
epithelial mucins. 9. Explain about formation of blood?
Ans : Blood : May be described as a specialised
connective tissue which circulates in a closed
system of blood vessels.
Blood Consists of

White blood cells Red blood cells Platelets Plasma

(WBC) (RBC)

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 MODEL PAPER -1 n 13

Function of blood : • Coagulation of blood: To stop bleeding by

• Respiration : Transport of oxygen from the the mechanism of clotting.
lungs to the tissues and carbon dioxide from Erythropoiesis: means the generation of eryth-
tissues to the lungs. rocytes. Upto the 3rd month of the intrauterine
• Excretion: Transport of metabolic wastes to life erythropoiesis occurs in the mesoderm of
the lungs, kidneys, skin and intestines for yolk sac. Between the 3rd month and the 5th
removal from the body. month of intrauterine life, erythropoiesis occurs
in the liver and spleen. This stage is called the
• Maintenance of normal acid base balance. ‘hepatic stage’ of erythropoiesis. From the 5th
• Nutrition: Transport of absorbed fatty ac- month, red bone marrow begins to produce
ids, monosaccharides and amino acids. erythrocytes.
• Regulation of water balance. Stages of Development: It has several stages.
• Regulation of body temperature. A pluripotent stem cell is the most primitive
cell, which gives rise to myeloid as well as
• Transport of hormones, vitamins and salts,
lymphocytic series cells.
which contain cations such as sodium, po-
tassium, calcium etc. and anions such as • The pluripotent stem cell then gives rise to
chlorides, phosphates, sulfates and carbon- committed stem cells. One type of commited
ates. stem cell produces RBCs, neutrophils, eosi-
nophils, monocytes, and platelets
• Transport of metabolites.
(b)Leukopoiesis: is the production of wbc. It
• Defence against infection by the white cells
has three indepedent series forms.
and the antibodies.

Granulocytes Lymphocytes Monocytes

In this series, the cytoplasm develops specific granules as the cell matures and the progressive
maturation of the nucleus is indicated by segmentation.
The stem cell or R.E. cell gives rise to other types of cells.
Development of Neutrophil: From the primitive precursor, pluripotent stem cell, develops the
committed stem cell and from this develops the progenitor cell, which is called colony forming
unit, granulocyte monocyte (CFU, GM). It is the common ancestor of both the neutrophil as well
as the monocyte. CFU, GM produces either the myeloblast (precursor of monocyte).
Myeloblast is a big sized cell (15 to 20m) with a very big nucleus, which almost completely fills up
the cell leaving a peripheral rim of cytoplasm. Nucleus contains 1 to 5 nucleoli and the cytoplasm
is nongranular. Promyeloblast, which forms later, resembles a myeloblast except that the cyto-
plasm appears pink coloured with nonspecific red granules. It contains one or two less defined
nucleoli and coarse chromatin.
Myelocyte formation takes place, which is smaller in size and contains round or oval nucleus with
larger pink cytoplasm. The nucleoli disappears and cytoplasm of this cell contains numerous
specific purple, red or blue coloured granules
Medical Lab Technician
14 n

Development of Eosinophil: From the pro- about 1000 platelets. Thus the platelets are
genitor cell, develops the eosinophilic my- produced in the red bone marrow and the
eloblast, which in turn develops into my- pluripotent stem, cell is the most primitive
elocyte and then into mature eosinophil. cell in the line of platelet production.
Development of Monocytes : Monoblast Thrombopoietin (TPO) is a polypeptide
is the earlier recognisable cell, which devel- responsible for the primary regulation of
ops in to the promonocyte and then into platelet production.
monocyte, then into monocytes are not seg-
It is produced by the liver, kidney, marrow
mented and ; are inserted in shape. The
stroma and other tissues.
cytoplasm is finely granulated. The cell size
is the largest of all the white blood cells seen • It stimulates the production and differentia-
in peripheral circulation. tion of megakaryotic precursor cells. it helps
for the full mutration of magakaryotic pre-
Development of Lymphocytes: The pre-
cursor cells. Thrombopoetin stimulates the
cursors of the lymphocyte: are present in
increase of the number of megakaryccytes
the red bone marrow. During the fetal life
of the bore marrow in the increase in num-
from red bone marrow some lymphocyte
ber of platelets.
precursors go to the thymus. There, they
are processed and become T-lymphocytes, 10. Write about collection of blood?
which are immunologically competent.
Ans : Methods of Collection of Blood:
These T lymphocytes go to the peripheral
Indications: Could be for one test or many
lymphoid tissue they make back and forth
tests related to various departments (pathol-
journey to blood.
ogy, biochemistry and microbiology). The
Some T lymphocytes become memory cells blood should preferably be collected early
within the lymph nodes. On receiving a fresh in the morning before the patient has eaten.
antgenic challenge.
Sites : From where blood can be collected.
Another precursor goes to the central lym-
I. Capillary Blood:
phoid tissue. From here, they move to the
peripheral lymph nodes and known as B 1. Finger tip of usually middle or ring finger.
lymphocytes. 2. Ear lobe.
3. Heal of sole - (done in children).
II. Venous blood : Ante cubital fossa.
III. Arterial Blood :
I. Capillary Blood: Skin Prick Method: Blood
may be obtained by pricking the skin of a
finger, the ear lobe or the feal of in an in-
fant.

c. Thrombopoiesis: The term thrombopoie- Instruments ® Needles with a flat body,

sis means generation of platelets. These are having a cutting edge or a sterile lancet or
developed from the giant cells called mega- No: 11 Bard parker blade are used.
karyocytes in the bone marrow. Procedure : The site is cleamed properly
• A single megakaryocyte can give rise to with alcohol or spirit, so as to kill bacteria
and also remove dirt and oils.
Medical Lab Technician
 MODEL PAPER -1 n 15

• Later the skin is allowed to dry Next, raise bow. These veins are easily seen and do not
the finger/heal, press the site between ball slip aside.
of index finger and thumb so as to form a
• The upper part of the arm is constricted us-
‘ridge’ then the lancet/needle is plunged into
ing a tourniquet or by placing B.P (blood
the ridge to a depth of about 3 mm and the
pressure) cuff and inflate upto 40 mmHg.
light pressure is immediately removed.
• Then, the patient is asked to clean the fist,
• The wound will antomatically open, and the,
or to open and close the fist for few times so
blood will starte to flow freely.
as to allow the veins to become prominent.
• The first drop of the blood is moped off,
• Next, identify and then feel the vein. Now,
and the next drop is used at once either for
clean the site with spirit or 95% acohol and
collection into a pipette or onto a slide, de-
allow the place to dry.
pending on the type of test needed. 2-3
blood semars may be made. • Next, place thumb of our left hand over the
vein just below the puncture site and apply
Once the required blood has been collected,
slight traction and ‘fix’ the vein.
the finger should be cleaned with a piece of
sterile cotton wool and pressure applied for • The syringe is taken in the right hand, and
30-40 seconds over the needle prick wound. bevel of the needle should face upwards,
and insert the needle at an angle of 30 de-
The capillary blood is used for the fol-
grees skin.
lowing tests:
• Push the needle firmly gently and teadily.
1. Estimation of Hb% and R.B.C., W.B.C and
Do not be fast, and avoid goind through
Platelet counts.
and through to come out from other side.
2. Preparation of peripheral smears.
• Once the needle is within the vein, pull the
3. Blood grouping. piston of the syringe, and the blood gets filled
4. Estimation of blood glucose level by shaffer into the barrel of the syringe.
somogyi’s micro-method • The correct required amount of blood is
2. Venipuncture : Amount of blood depends withdrawn, and the tourniquet is removed.
on the number and type of tests required. • Syringe along with the needle is smoothly
And based on this,the proper size of syringe and gently removed.
(2ml, 5ml or 10ml) is used. The syringe
• Place a sterile guaze piece over the punc-
should be dry and sterile, if not dry, the red
ture site and pressure is applied by flexing
blood cells (RBC) will lyse.
the arm for several minutes, which prevents
Needles used for venipuncture are about 1- bleeding by closure of the wound.
1 1/2 inches long with a medium size bore
• The blood which is di awn, should be im-
of about 18-20 guage. The tip of the needle
mediately transferred to the appropriate con-
should be sharp. The needle should fit into
tainers as per the tests needed.
the syringe properly and should not allow
air inside or blood to leak out. • This is done by first removing the needle
from the syringe and the transfering blood
Procedure : The patient is made to sit down
into different containers.
with the fore arm resting on a table to or lie
down with the arm resting. Venous blood is necessary for the following
tests:
• One of the veins in the tante aute cubital
fossa is chosen which is in front of the el- • Estimation of E.S.R. P.C.V. etc.
Medical Lab Technician
16 n
• Estimation of blood constituents like sugar,
11. Write in detail regarding blood banking
urea, etc.
Ans : Blood collection is the most important func-
• Bacteriological and serological examinations.
tion of the blood transfusion centre. Donors
• Blood grouping and cross matching. should be healthy and strict criteria for do-
III. Arterial Blood: nor selection must be followed.

• Sometimes it may be impossible to collect Types of donors

blood from veins. In such cases arterial Rapid commerical donors (PCD)
puncture can be attempted, Brachial artery
and radial artery are the usual sites. Replacement donors (RD)

• Sometimes arterial blood gives positive cul- Voluntary donors (VD)

ture results, when venous blood is negative 1. P.C.D : They are professional blood donors
in case of subacute bacterial endocarditis. (blood sellers) who receive monetary ben-
Containers for different tests: The contain- efit for donation of blood. These donors are
ers vary, depending on the types of tests usually not healthy and are anemic and at
necessary to be carried out on the venous risk of transfusion, transmissible infection.
blood obtained. 2. Replacement Donors : Individuals who are
• For Haematology tests: Usually ‘penicillin’ self motivated or friends of a patient or family
bottles are used, with appropriate amount members and donate blood for their rela-
of anticongulants used for that particular tive in need.
test. 3. Voluntary Donors : Individuals who are self
• Now a days, vacutainers are used, which can motivated and come forward to donate
be directly centrifuged without transferring blood with the prime aim of helping on un-
into another container. known recipient. They do not desire for any
monetary personal benefits.
Blood For Biochemical and serological
tests: Blood is usually collected in a dry, Procedure: This should be carried in the
sterile test tube. No anticoagulant is added, presence of a Medical officer, Identification
as most biochemical and serological tests are of donor by donor number is necessary.
done an the serum. Emergency ‘Kit consisting of intravenous flu-
• Serum is obtained by allowing the col- ids, adrenaline, dexamethasone, calcium
lected blood to stand undisturbed for gluconate, oxygen cylinder with regulator
some time. and mask and Mephentermine, is a must at
every centre where phlebotomy is done.
• Blood clot forms and retracts, and the
serum separates. The area in the antecubital fossa is cleaned
and the vein is selected and a sphygmoma-
• This serum can be removed with the help
nometer cuff is applied and pressure is raised
of a sterile pasteur pipette.
to 60 mm of hg and insert the needle at
Blood for Bacteriological tests: Usually strin- 45°C with bevel upwards into the vein.
gent aseptic precautions are taken, when blood
is collected for bacteriological tests, espeically, Ask the donor to make a clean first and re-
when it is done for culture.Approximate sterile lease it during collection of blood. See that
receptacles containing various culture media are there is no air in the bag for collection of
used which will facilities the growth and helps blood.
in isolation and identification of the organisms.
Medical Lab Technician
 MODEL PAPER -1 n 17

Mix the blood and anticoagulant gently and • In the past 1 month before donating did
periodically during collection of blood. the donor had any extensive dental
work.
• When the appropriate amount has been col-
lected, clamp the tubing with artery forceps • Was he exposed to any infection.
and deflate the cuff. • Skin rash or infection.
• Normally 420 ml of blood is collected in a • Suffering from Cold, sore throat, asthma,
big bag which has 80 ml of acid citrate dex- active allergy etc.,
trose (ASD) as anticoagulant.
• Did the donor have a positive test for
• Place the sterile swab at the site of juncture AIDS or Hepatitis B.
after completion and with drawing of the
• For women - H/o Pregnancy in the last
needle.
12 months.
• Post donation Donor care Make the donor
• No. of previous donations and the date
lie down comfortably and he should be
of last donations.
down on the couch for a short while upto
3-5 min after phlebotomy encourage the After clear information the following
candidate to have more fluids and to avoid testtube done.
smoking, alcohol, and strenuous exercise for
1. HB estimation. 2. Weight.
24 hrs.
3. B.P. 4. Veneral disease.
Note: Nearly everybody can donate a 500
ml unit , a donor can donate at one time 5. Blood group. 6. Test for hepatitisB.
wihtout trouble. 7. HIV (AIDS). 8. Hepatitis C.
The donor should be examined for and 12. Write the following.
details should be elicited of:
1. Anticoagulants
• Good health.
2. Blood cell count.
• H/o. Epilepsy of convulsions.
Ans. Anticoagulants : The blood has to be kept
• H/o Heart disease. in the fluid state for many of the haemato-
• H/o Tuberculosis. logical and bio chemical examination.
• Incidences of bleeding tendency. • In order to achieve this, anticoagulants have
to be added in an appropriate porportions.
• Diabetes.
• Following are the various anticoagulants
• H/o Janudice.
commonly used.
• Did the donar receive blood transfusion
1. Oxalates : These salts unite with calcium
in the last 12 months period.
in the blood to form insoluble com-
• Any immunisation or vaccination. pounds and thereby deplete the blood
of its calcium which is necessary for the
• In the past 6 months, did the donor un-
coagulation of blood.
dergo any serious injury.
a. Dried Potassium Oxalate : Is used at a
• Any surgery
concentration of 2mg /ml of blood. Dis-
• In the past 6 months did the donor had advantages causes shrinkage and de-
malaria. struction of cells. It is often used for
chemical analysis of blood.
Medical Lab Technician
18 n

b. Ammonium oxalate : Used in case of rocytes are better preserved in the ACD than
estimation of blood constituents. 2 mg. in Trisodium Citrate alone.
of dried salt is sufficient for 1 ml of blood. • Tri-sodium citrate : 1.32 g
Disadvantage : It causes swelling of RBC • Citric acid : 0.42 g.
therefore, not used for PCV, ESR or pe-
ripheral smear. • Dexttrose : 1.40g

c. Double oxalate or mntrobe’s salt: Com- • Dist. water add to : 100 ml .

bination of ammonium and potassium 15 ml. of ACD is sufficient for 100 ml. of
oxalates (1.2g and 0.8 gm of these salts blood
respectively and distilled water of 100 ml)
is to be used. Uses : This is used in blood transfusion.

• There are 2 mg in 0.1 of the solution. 3. Heparin : It has an affinity for blood pro-
teins and acts as an antithrombin and antith-
• The fluidis evaporated and dried mix- romboplastin, lmg. of dry Heparin or 1000
ture is to be used. This solution contains I.U. of liquid Heparin suffice for 10 ml. of
20mg/cc. blood.
Advantages: • Used in Haematocrit studies and blood
• Ammonium oxalate causes swelling of the transfusion especially exchange and rapid
cells, whereas potassium oxalate shrinks. transfusion as in cases of thoracic surgery,
E.S.R. and osmotic fragility tests.
• The action of both these salts is counter
balanced and thereby the cells retain in 4. Ethylene Diamine Tetra Acetate (EDTA):
original shape and size. • Its disodium and dipotassium salts are used.
Uses: It is the most powerful chelating agent and
thus separate the calcium ions from the
• Used in the estimation of E.S.R. by blood.
Wintrobe’s method and P.V.C., HB esti-
mation, WBC, RBC, prothrombin time • Used for most of the the hamatological and
etc. chemical tests

• Not used for peripheral blood smears, Blood Cell Counts: The technique of count-
because leukocytes often phagocytose ing of the blood cells is known as hemocyto-
the calcium oxalate precipitate. metry. This involves manual counting of the
cells with the help of a microscope after dilut-
2. Citrates: ing blood in respective diluting fluids. The cell
a. Trisodium citrate: most often counted by this techniques are red
cell, white V cells, platelets and eosinophils. The
• Part of 3.8% solution of thesalt and 9 parts hemocytometer technique, however, can not
of blood mixed for coagulation studies. differentiate the various types of the individual
• Used in blood transfusion as the salts are cells.
relatively nontoxic and the salt is excreted Requirements:
by the kidneys or utilised by the body.
1. Microscope.
• 3.8% solution is used as 1:4 ratio with blood
for ESR estimation (Westergren’s method). 2. RBC Pipette.
b. Acid Citrate Dextrose Soln. (ACD): Eryth- • This pipette has a large bulb, which con-

Medical Lab Technician

 MODEL PAPER -1 n 19

Important Precautions: The precautions must be

tains a red glass bead. It has three marks
discarded and the filling procedure should
0.5, 1.0 and 101.
be repeated
• Blood is drawn to 0.5 mark and then
by using another clean and dry chamber if:
diluting fluid is drawn up to the mark 101.
The dilution of blood is 1: 200. 1. Chamber area is incompletely filled.
3. WBC Pipette. 2. Fluid overflows into moat.
• It is smaller than the red blood cell pi- 3. Air bubbles are seen anywhere in chamber area.
pette and its bulb contains a white glass 4. Any debris is seen in chamber area.
bead.
• It has three marks 0.5, 1.0 and 11.
• The blood is drwn up to 0.5 mark and
then diluting fluid is drawn up to 11
mark. The dilution of blood is 1:20.
4. The counting Chamber
• The chamber most commonly used is the
improved Neubaur chamber which has
an area of 9 sq. mm and a depth of 0.1
mm.
• The two stages are seperated from two
ridges, one on either side, by a gutter.
• The surface of the two ridges is 1/10 mm
above the stage.
• The 9 sq. mm area of the counting cham-
ber is divided into 9 squares. The four
corner squares measure 1 sq. mm area.
• The central square is divided into 25
squares and each of these square is fur-
ther divided into 16 small squares, each
square measures 1/400 sq. mm in area.
• Five of the medium squares (or 80 small
squares) are used in counting red blood
cells.
5. Cover slips: Aspecial type of cover slip is used
with a very smooth and even surface. These
are available in two sizes
a. 16 × 22mm -
b. 22 × 23mm -
The thickness of the cover slips may be 0.3
mm, 0.4 mm or 0.5 mm.

Medical Lab Technician