Tissue culture involves the use of small pieces of plant tissue (explants) which are cultured in a nutrient medium under sterile conditions. Tissue culture is in vitro maintenance and propagation of isolated cells tissues or organs in an appropriate artificial environment.

1. 1
Plant Tissue Culture
Sl no Name of the experiment Page no Date of
experiment
1 M.S Media preparation and sterilization 03
2 Preparation of explants and callus induction 06
3 Sub culture and maintenance of callus 08
4 Regeneration of plant from callus 09
5 Meristem culture for pathogen free plants 10
6 Suspension Culture 11
7 Synthetic seeds 12
8 Preparation of PDA 13
Animal Tissue Culture
Sl no Name of the experiment Page no Date of
experiment
1 Preparation of Hank’s balance salt solution 14
2 Cell viability test using Trepan blue exclusion method 15
3 Observation of poly Morpho Nuclear Leucocytes 17

2. 2
Experiment no 1
M.S Media preparation and sterilization
AIM: Preparation of stock solutions of MS (Murashige & Skoog, 1962) basal medium and plant growth
regulator stocks.
PRINCIPLE:
The basal medium is formulated so that it provides all of the compounds needed for plant growth and
development, including certain compounds that can be made by an intact plant, but not by an isolated piece of
plant tissue. The tissue culture medium consists of 95% water, macro- and micronutrients, vitamins, amino
acids, sugars. The nutrients in the media are used by the plant cells as building blocks for the synthesis of
organic molecules, or as catalysators in enzymatic reactions. The macronutrients are required in millimolar
(mM) quantities while micronutrients are needed in much lower (micromolar, µM) concentrations. Vitamins are
organic substances that are parts of enzymes or cofactors for essential metabolic functions. Sugar is essential for
in vitro growth and development as most plant cultures are unable to photosynthesize effectively for a variety of
reasons. Murashige & Skoog (1962) medium (MS) is the most suitable and commonly used basic tissue culture
medium for plant regeneration. Plant growth regulators (PGRs) at a very low concentration (0.1 to 100 µM)
regulate the initiation and development of shoots and roots on explants on semisolid or in liquid medium
cultures. The auxins and cytokinins are the two most important classes of PGRs used in tissue culture. The
relative effects of auxin and cytokinin ratio determine the morphogenesis of cultured tissues.
MATERIALS:
• Beakers (100 ml, 500 ml and 1000 ml)
• Measuring cylinders (500 ml)
• Disposable syringes (5 ml)
• Disposable syringe filter (0.22 µm)
• Autoclaved eppendorf tubes (2 ml)
• Eppendorf stand
• Benzyl-aminopurine
• Naphthalene acetic
Media constituents
Inorganic nutrients
Mineral elements are very important in the life of a plant. Besides, C, H, N, and O, 12 other elements are known
to be essential for plant growth. According to the recommendations of the International Association for Plant
Physiology, the elements required by plants in concentration greater than 0.5 mmol/l are referred to as
macroelemetns or major elements and those required in concentration less than the prescribed amount are
microelements of minor elements. A variety of salts supply the needed macro and micronutrients that are the
same as those required by the normal plant.
Major salts : The salts of potassium (K), nitrogen (N), calcium (Ca), magnesium (Mg),phosphorus (P) and
sulphur (S) are required in macro or millimole quantities. Nitrogen is generally used as nitrate or ammonium
salts, sulphur as sulphates and phosphorus as phosphates.
Minor salts : The salts of iron (Fe), manganese (Mn), boron (B), copper (Cu), zinc (Zn), iodine (I), molybdenum
(Mo) and cobalt (Co) are required in micromolar concentrations and are considered to be minor salts. These
salts are essential for the growth of tissues and are required in trace quantities.
Carbon and energy source
The standard carbon source without exception is sucrose but plant tissues can utilize a variety of carbohydrates
such as glucose, fructose, lactose, maltose, galactose and starch. In the cultured tissues or cells, photosynthesis

3. 3
is inhibited and thus carbohydrates are needed for tissue growth in the medium. Sucrose, at a concentration of
2-5% in the medium, is widely used. The autoclaving process does cause an alteration in the sugars by
hydrolysis but presents no drawbacks to the growth plan. Most media contain myoinositol at a concentration of
100-mg per litre, which improves cell growth.
VITAMINS
Normal plants synthesize the vitamins required for growth and development, but plant cells in culture have an
absolute requirement for vitamin B1 (thiamine), vitamin B (nicotinic acid) and vitamin B6 (pyridoxine). Some
media contain pantothenic acid, biotin, folic acid, p-amino benzoic acid, choline chloride, riboflavine and
ascorbic acid. The concentrations are in the order of one mg/l. Myo-inositol is another vitamin used in
the nutrient medium with a concentration of the order of 10-100 mg/l.
GROWTH REGULATORS
Hormones now referred to as growth regulators are organic compounds that have been naturally synthesized in
higher plants, which influence growth and development. These are usually active at different sites from where
they are produced and are only present and active in very small quantities. Two main classes of growth
regulators of special importance in plant tissue culture are the auxins and cytokinins, while others are of minor
importance, viz., gibberellins, abscisic acid, ethylene, etc. Some of the naturally-occurring growth regulators are
indole acetic acid (IAA), an auxin and zeatin and isopentenyl adenine (2 iP) as cytokinins, while others are
synthetic growth regulators.
Auxins
Auxin was discovered following experiments on the reaction of coleoptiles curvature in Gramineae. It owes its
name to its effect on the elongation of cells (auxesis).
Auxins have an indole nucleus with the basic formula C10H9O2N.A common feature of auxins is their property
to induce cell division and cell elongation. The stimulation of division of cells of cambial origin resulted in
initial successes with in vitro cultures. This effect leads to a number of cells, which further result in the
formation of callus. Auxin has a clear rhizogenic action, i.e. induction ofadventitious roots. It often inhibits
adventitious and auxillary shoot formation. At low auxin concentration, adventitious root formation
predominates, whereas at high auxin concentration, root formation fails to occur and callus formation takes
place. All the plants synthesize auxin that is modulated according to the stage of development. Auxin is present
in sufficient concentration in the growing shoot tips or flowering tips of plants to ensure cell multiplication and
elongation. Auxin circulates from the top towards the base of the organs with a polarity strongly marked in
young organs. The compounds most frequently used and highly effective are 2,4-dichlorophenoxy acetic acid
(2,4-D), naphthalene acetic acid (NAA), indole acetic acid (IAA), indole butyric acid (IBA).Other auxins in use
are 2,4,5-trichlorophenoxy acetic acid (2,4,5-T), p-chlorophenoxy aceticacid (pCPA) and pichoram (4-amino-
3,5,6-trichloropicolinic acid).
Cytokinins
Cytokinins were discovered during in vitro culture studies. Coconut milk was known to have a favourable effect
on cellular multiplication and bud formation. Cytokinins are derivatives of adenine and have an important role
in shoot induction. Cytokinins also have a clear effect on cell division. Often used to stimulate growth and
development, they usually promote cell division if added together with an auxin. Auxins favour DNA
duplication and cytokinins enable the separation of chromosomes. Cytokinins have a clear role in organogenesis
where they stimulate bud formation. They are antagonistic to rhizogenesis. At higher concentrations (1 to 10
mg/l), adventitious shoot formation is induced but root formation is generally inhibited. Cytokinins promote
axillary shoot formation by decreasing apical dominance. The most frequently used compounds are kinetic,
benzyl adenine (BA) or 6-benzyl amino purine (BAP), zeatin, and isopentenyladenine (2 iP). Zeatin and 2 iP are
natural cytokinins.

4. 4
Gibberellin
Gibberellins are normally used in plant regeneration. GA3 is essential for meristem culture of some species. In
general, gibberellins induce elongation of internodes and the growth of meristems or buds in vitro. In its
absence, the culture appears globular, due to the accumulation of nodes. Gibberellins usually inhibit
adventitious root as well as shoot formation. During organogenesis, gibberellins are antagonistic. They seem to
oppose the phenomenon of dedifferentiation. Thus, in in vitro cultures, they cannot be used for this purpose, but
can be utilized for explants already organized (meristems, apices, buds).
Preparation of M.S media
 An appropriate amount of stock solution of salt are transferred to one liter flask
 To this organic suppliments ( coconut milk , malt)carbon source(glucose or sucrose) are added
 The final volume is made up to liter by adding distilled water and the pH is adjusted to 5.0 -5.8
 Solidifying agent agar is added to get semisolid media
 The melted medium is poured in to culture vessels and plugged with non absorbent cotton and sterilized
at 121 ºC for 20 min at 15lbs
 Media is allowed to cool and stored at 25 ºC for further use.
MS NUTRIENTS STOCKS
Nutrient salts and vitamins are prepared as stock solutions (20X or 200 X concentrations of that required in the
medium) as specified. The stocks are stored at 4
0
C. The desired amount of concentrated stocks is mixed to
prepare 1 liter of medium.
Murashige T & Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures.
MS major salts mg/1 L medium 500 ml stock (20X)
1. NH4
NO3
1650 mg 16.5 gm
2. KNO3
1900 mg 19 gm
3. Cacl
2
.2H
2
O 440 mg 4.4 gm
4. MgSO4
.7H2
O 370 mg 3.7 gm
5. KH2
PO4
170 mg 1.7 gm
MS minor salts mg/1 L medium 500 ml stock (200X)
1. H3
BO3
6.2 mg 620 mg
2. MnSO4
.4H2
O 22.3 mg 2230 mg
3. ZnSO4.
4H2
O 8.6 mg 860 mg
4. KI 0.83 mg 83 mg
5. Na2
MoO4.
2H2
O 0.25 mg 25 mg
6. CoCl
2.
6H
2
O 0.025 mg 2.5 mg
7. CuSO4.
5H2
O 0.025 mg 2.5 mg

5. 5
MS Vitamins mg/1 L medium 500 ml stock (200X)
1. Thiamine (HCl) 0.1 mg 10 mg
2. Niacine 0.5 mg 50 mg
3. Glycine 2.0 mg 200 mg
4. Pyrodoxine (HCl) 0.5 mg 50 mg
Iron, 500ml Stock (200X)
Dissolve 3.725gm of Na
2
EDTA (Ethylenediaminetetra acetic acid, disodium salt) in 250ml dH
2
O.
Dissolve 2.785gm of FeSO4
.7H2
O in 250 ml dH2
O
Boil Na2
EDTA solution and add to it, FeSO4
solution gently by stirring.
PLANT GROWTH REGULATOR STOCK
The heat-labile plant growth regulators are filtered through a bacteria-proof membrane (0.22 μm) filter and
added to the autoclaved medium after it has cooled enough (less than 600
C). The stocks of plant growth
regulators are prepared as mentioned below.
Plant Growth
Regulator
Nature Mol. Wt. Stock
(1 mM)
Soluble in
Benzyl
aminopurine
Autoclavable 225.2 mg/ ml 1N NaOH
Naphtalene
acetic acid
Heat labile 186.2 mg/ ml Ethanol
Experiment no 2
Preparation of explants and callus induction
Aim: choose explants and prepare for inoculation and induction of callus
Plant preparation
Principle : Plant cells and tissues are totipotent in nature i.e., every individual plant cell or tissue has the same
genetic makeup and capable of developing along a "programmed" pathway leading to the formation of an entire
plant that is identical to the plant from which it was derived. The totipotency of the plant cells and tissues form
the basis for in vitro cloning i.e., generation or multiplication of genetically identical plants in in vitro culture.
The ability to propagate new plants from a cells or tissues of parent plant has many interesting possibilities.
Requirements :Beakers, Measuring cylinders, Conical flasks, Cotton plugs, Sucrose, BAP (1mM stock), Agar
Agar, Forceps, Blade Holder, Sterilized blades , NAA (1 mM stock), Micropipettes, sterilized microtips, cork
borers, petridishes.
Procedure
 Any living part of the plant used for culturing in tissue culture is called as explant.
 The explant of a garden or potted plant carries germs on its surface. Hence its surface must be cleaned by a
detergent in running water and surface sterilized with disinfectants like Sodium hypochlorite solution.
 Later rinsed in distilled water.
 Hands are sterilized with 70% alcohol and allowed to dry
 Trim the explants to different size

6. 6
 The sized explants are transferred to petriplate containing 0.1% mercuric chloride
 Then wash it with water to remove trace amounts of mercuric chloride
 The mouth of the culture tube is flamed and the explants are aseptically transferred to the culture tube
containing sterilized media.
 The mouth of the tube is again and the mouth of the tube is again plugged immediately.
 Then the tubes are incubated at 25ºC under16 hr of photoperiod with ~1000lux light intensity for 1 to 2
weeks.
 Record the observation

7. 7
Experiment no 3
Sub culture and maintenance of callus
Aim: To maintain the subculture and maintain callus culture.
Principle: Growing culture exhausts the media in culture flasks, when the nutrient level comes down the callus
is transferred to fresh medium, the maintenance of fragmented callus in fresh nutrient medium is called
subculture.
Requirement: MS media supplemented with 2,4,D in culture flasks, forceps, petriplate, laminar air flow unit.
Procedure:
 Subculture of the callus is carried out under aseptic condition in laminar air flow.
 With the help of sterile process soft callus is broken down in to small pieces.
 The smaller portion of callus is transferred to fresh media and flasks are incubated under light at 25+
2ºC.
 Culture flasks are periodically checked for contamination and proliferation or growth of callus.
 Generally subculture is done at regular intervals of about 4 weeks.
Culture tube cut pieces of callus Callus transferred to fresh culture tubes
Containing callus
Report – Callus growth is maintained further on sub culturing with the same or higher alcohol concentration.

8. 8
Experiment no 4
Regeneration of plant from callus
Aim: to regenerate callus using growth regulators
Principle: The M.S medium is most suitable for plant regeneration from tissue and callus. The hormones are
the most important components in plant regenerating media. The capacity for plant regenerating tissue, vary
widely in different species. In some species the morphogenesis is readily induced (carrot, coffee)and developed
in to complete plant while in others fails to occur.
Requirements: Callus, M.S liquid medium, shaker, auxins , cytokines, forceps, pots, green house, growth
chamber etc
Preparation of rooting medium-
M.S rooting media is formulated with high proportion of auxin
Ex- 1) IAA (3mg/lt) and low proportion of cytokines.
2) BAP or kinetin (0.02mg/lt) induce root developed from callus
Procedure
 Transfer callus in to flask, each containing 20 ml of liquid M.S media
 Shake the culture flask on a shaker at 150rpm to disassociate callus in to single cells
 Aseptically transfer a small callus (tissue) on to M.S medium with 3mg/lt Indole acetic acid and 0.02
mg/lt of BAP
 Incubate at 25 to 27ºC in light (100klux) to develop the tissue in to a large callus and shoot for four to
five weeks.
 When shoot appears transplant these on M.S medium with 0.03mg/lt IAA and 2mg/lt BAP.
 Keep the pots in green house or growth chamber where high humidity is maintained and then planted in
main fields.
Observation- appearance of callus and shoot takes place after four weeks of incubation on M.S media
supplemented with hormones. Root appears later from the callus with shoots in M.S rooting media.
The plants in the plant will grow to maturity flower and set seeds similar to the plants that grow in nature.

9. 9
Experiment no 6
Meristem culture for pathogen free plants
Aim: To produce pathogen free potato by meristem tip culture
Principle: Shoot meristem culture is a technique in which a dome shaped portion of the meristem region of
the stem tip is dissected and inoculated into nutrient media which supports growth. The dome region is
approximately 0.075-0.12mm 0.22 - 0.25mm in size. The apical dome is not connected to procambium
though vascular system or to young leaf primordial on young axillary buds this lack of vascular connection
provides a basis for using the meristem technique for pathogen elimination .
Requirements;
Scalpel, potato, 0.1% HgCl2, nutrient media, forceps, 70% alcohol, incubation facilities
Procedure:
 The tubers were surface sterilized with 0.1% HgCl2 followed by sterile distilled water wash
 The tubers were cut at the eye region
 The meristem explants were transferred on to M.S media with BAP.
 The culture were incubate at 25ºC with 12hr light period
Surface sterilised tuber Eyes of potato Explant induction Plant let
. in to media regeneration
Result:

10. 10
Experiment no 7
Suspension Culture
Aim: To initiate suspension culture from callus.
Principle: unorganised plant calls can be cultured as callus and also as cell suspension culture in agitating
liquid medium.
The fast growing fragile callus is used for initiating callus
suspension culture. A medium used for initiating species is
suitable for suspension culture. The medium is enriched with
growth hormones . The cell suspension is kept agitated for
aeration, uniform distribution and to get free cells during
incubation period biomass increases due to cell enlargement and
cell division. Suspension cultures are widely used as model system
for studying developing pure line of cells for cell hybridization,
gene expression and for extraction of secondary metabolites.
Requirement: callus, M.S liquid medium, 2, 4-D, 70% alcohol,
Forceps, laminar air flow unit.
Procedure:
 The standard protocol for laminar air flow unit sterilization
along with inoculation set work is carried out
 Callus of the plant grown on solid M.S media were removed
on to a sterile petriplate.
 With the help of sterile forceps and scalpels the callus were
cut into small pieces.
 The calluses were then transferred in to conical flasks
containing liquid M.S media.
 The conical flasks were then incubated at 23+2ºC On a rotating shaker.
 To subculture, an aliquot of the growing cell suspension was transferred to fresh media using a pipette.
Result:

11. 11
Experiment no 8
Synthetic seeds
Aim: To prepare synthetic seeds by immobilization technique using Ragi seeds.
Principle: synthetic seeds are defined as somatic embryos engineering in commercial propagation of plant, they
are by the a technique of immobilization of somatic embryos are encapsulated with alginate which forms a semi
permeable through which water and nutrients are taken up from media.
Immobilised or synthetic seeds can be handled mechanically, easy to storage and transport.
Materials Required:
Beaker
Petri dish
Micropipette
Microtips
Sodium alginate (4%)
Calcium chloride (4%)
Distilled water
Procedure:
1. Sodium alginate (4%) and Calcium chloride (4%) are prepared and autoclaved
2. M.S basel solid media is prepared and autoclaved
3. The selected seeds are surface sterilized with 1% MgCl2 and washed few times with sterile distilled water.
4. It was transferred to sodium alginate solution and incubated it for 5- 10 mins.
5. The seeds were then dropped carefully from sodium alginate to chilled calcium chloride solution in such a
way that each drop contains a single seed.
6. The beads formed were transferred to a separate plate and washed in sterile distilled water and incubated on
solid M.S media.
7. Incubated at23+2ºC for almost two weeks the media is observed for germination.

12. 12
Result:
Hydrated or encapsulated seeds are formed and germination of the seeds was observed after_____ days.
Preparation of PDA

13. 13
Animal Tissue Culture
Experiment no 1
Preparation of Hank’s balance salt solution
Introduction
 A basal salt solution us composed if inorganic salts, glucose,BSA, bicarbonates etc,
 This basal salt solution provide water and certain ions for normal cell metabolism while maintaining
intracellular and extra cellular osmotic balance which will also provide a buffering system to maintain
the physiological pH
Procedure:
1. Solution A: Dissolve 5gm of dextrose ,300mg of KH2PO4 and 375mg of NAH PO4,2H2O in 250 ml of
distilled water
2. Solution B: Dissolve 40gm of sodium chloride ,2gm of KCl , 0.5gm MgSO4.7H2O,0.5gm of
magnesium chloride hexahydrate,0.977gms of calcium chloride in distilled water and make the volume
to 250 ml
3. Working solution- Dissolve solution A and solution B in 1:10 propotion and add BSA at concentration
of 0.4%when ever required.
Application
 Used to Isolation of lymphocytes for immunological preparations, DNA damage study
 Used to maintain viability of cells and as a good suspension medium
 Used As a diluting fluid in flow cytometry of animal cells.

14. 14
Experiment no 2
CELL COUNTING AND VIABILITY
Aim: To ensure the population of cells required for the culture works by cell counting method and its
Viability by vital staining methods
Introduction
Haemocytometer (also known as hemocytometer) is a glass slide with two counting chambers etched
in a surface area of 9mm square. Each chamber is divided into nine 1.0mm square. It has raised sides which
keep the cover slip 0.1mm above the chamber floor so that the total volume of each square becomes
0.0001ml(1.0mm x 0.1mm or 0.1mm² or 10² cm³, L x W x H ).
Principle
Staining of cells identifies viable cells. Stains generally used are Trypan Blue, Erythrosin B and
Nigrosin. Nuclei of damaged or dead cells take up the stain whereas the viable cells do not do so.
Requirements
Cell suspension
Spirit lamp
Hemocytometer
Microscope
Micropipette
Tryphan Blue 0.4%
Procedure:
1. Take the Hemocytometer and place it on the flat surface of the work bench. Place the cover slip on the
counting chamber.
2. Mix 20μl cells that have been well mixed prior to sampling with an equal volume of tryphan blue.
3. Apply to a hemocytometer by pipetting from the edge of the cover slip and permitting diffusion by
capillary action.
4. Make sure that there is no air bubble and there is no overfilling beyond the ruled area.
5. Leave the counting chamber on the bench for 2-3 minutes to allow the cells to settle.
6. Place the counting chamber on the stage of the microscope between the clips to the hold slide so that the
counting chamber can be moved (if the microscope is provided with a moving stage).
7. Switch to low power (10x) objective, adjust the light (less light needed, hence close the aperture or lower
the condenser) and focus on the wall of the counting chamber.
8. Then slowly move the stage towards the middle of the slide until the ruling area visible, sharpen the focus
and locate the large square in the centre.
9. Locate the large square in the centre with 25 small squares. Place in the middle of the field of vision and
examine the distribution of viable cells on the entire area. It must be uniform or else refill the chamber with
cell suspension.
10. Carefully switch to high power objective (40 x) and move the chamber so that the smaller upper left corner
square (with 16 smaller squares) is completely in the field of vision.
11. Count the number of unstained cells seen on the small square (0.2x0.2=0.04sq mm) of the upper left
corner which is divided into 16 smaller squares to facilitate counting.
12. Repeat the counting with three other corner squares.
13. Make a total of all the cells counted in 4 squares.

15. 15
Result
The percentage of viable cells = ________
The percentage of non viable cells = ________
Observation and calculation:
Squares
counted
Viable cell
count (W)
Non viable
cell count (Y)
Total cell
counted (Z)
A
B
C
D
Total
1. Total number of cells /ml= Z ×25×104
cells/cm3
=
=
2. Total number of viable cells /ml= W ×25×104
cells/cm3
=
=
3. Total number of non viable cells /ml= Y ×25×104
cells/cm3
=
=
Percentage of viable cells = W ×100
Z
Percentage of non viable cells = Y ×100
Z

16. 16
Experiment no 3
Observation of Poly Morpho Nuclear Leucocytes
Granulocytes: (Poly morpho nulearleucocytes)
Leucocytes are characterised by the presence of differentially stained granulocytes in this cytoplasm when
viewed under light microscope. These granules are membrane bound organs which primarily act in digestion
and endocytosis of particles. There are 3 types of granulocytes they are, neutrophiles, basophiles and
eosinophiles which are named according to their staining property.
Neutrophiles:
 Diameter : 10-12 µmts
 Nucleus : Multilobed nucleus
 Granules: fine, faintly pink in E and H stain
 Lifetime : 6 hrs to few days
 %in adult blood: 60-70%
 Functions: mainly targets bacteria fungi and other inflammatory process. They are first response against
microbial infections their activity, death in large number forms pus.
Eosiniphiles:
 Diameter : 10-12 µmts
 Nucleus : Bilobed nucleus
 Granules: Full pink or orange in E and H stain
 Lifetime : 8 – 12 days
 %in adult blood: 1-6%
 Functions: preliminary act against large parasitic infections and also in inflammatory and allergic
reactions.
Basophiles
 Diameter : 12-15 µmts
 Nucleus : Bilobed or trilobed nucleus
 Granules: Large, blue
 Lifetime : Few hrs to few days
 %in adult blood: Less than 1%
 Functions: Chiefly responsible for allergic and antigen antibody reaction by release of histamines.
Agranulocytes (mononuclear leucocytes)
Leucocytes characterized by absence of granules in their cytoplasm. These cells do contain non specific
granules like lysosomes.
There are two types of agranulocytes they are lymphocytes and monocytes.
Lymphocytes:
 Diameter : 7-8 µmts
 Nucleus : Deeply stained eccentric nucleus
 Granules: absent
 Lifetime : Weeks to years
 %in adult blood: 25-33%
 Functions:
i. Activation of B cells producing antibodies against antigen and causes B cell memory.

17. 17
ii. Activation of T cells against viral infections and in turn activates natural killer cells and
involve in a wide range of cell destroying activities.
Monocytes:
 Diameter : 14-17 µmts
 Nucleus : Kidney nucleus
 Granules: Absent
 Lifetime : hrs to days
 %in adult blood: 2-10%
 Functions: Monocytes have phagocytosing activity, it phagocytes pathogens and antigens and process
them and present them to T cells. They can be differentiated in to macrophages and dendritic cells.
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