Download MBBS Important Topics Micro Practical

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Slide

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1 drop normal saline

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a loop of bact. Colony.

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air day

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Heat fix

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methylene blue/ Crystal violet/ gentian violet for 1 min.

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Iodine 1 min

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wash with water

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Acetone/ alcohol 2-3 sec.

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wash

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Conterstain carbol fuschin/ neutral red/

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Safranin 1-2 min

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wash → dry → oil immersion → 100%

Principle

• Primary stain (basic) + Iodine → insoluble complex in cytoplasm

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• decolorrises dissociate this compound and remove from cell. This slower in G^+ve
• Reason:- (1) G^+ve have thicker & dense peptidoglycan cell wall which is less permeable
• (2) G^-ve have acidic cytoplasm so bind more firmly with basic dye.

G^+ve look like G^+ve when → overdecolorrigation

• meerupture of cell wall during heat fixation

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G^+ve look like G^+ve when → under decolourisation

G^+ve cocci (violet, rounded) → G^+ve bacilli (violet, rod)

• Staphylococci → M. tuberculosis
• Streptococci → M. leprae
• Micrococci → Clostridium

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• Pneumococci → corny bactarium
• enterococci → Bacillus Anthrasis

G^-ve cocci (pink, round) → G^-ve bacilli (Pink, rod)

• Neisseria meningitidis → E. coli
• Neisseria gonorrhoea → Proteus

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• moraxella → Pseudomonas
• Salmonella
• Shigella

Crystal violet solⁿ = Crystal violet (0.5gm)+ dist. water (100ml)

Gram's iodine = Iodine (1gm) + KI(2gm) + dist. water (100ml)

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Safranine = Safranine (0.5gm) + dist. water (100ml)

AF +ve → M. tuberculosis

↓ M. leprae (need 5% H₂SO₄)

Nocardia (need 1% H₂SO₄)

Heat fixed slide

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Brick red bacilli with blue background.

Carbol fuschin (blue coloured pus cells may be seen)

Spore Head of sperm

Heat till steam arises

Principle :- mycolic acid (& Hydroxy, long chain fatty acid) present in cell wall so heat is applied for penetration of carbol fruschin and mycolic acid is also responsible for resist to decolorrisor.

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wait 5 min.

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wash

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20% H₂SO₄ 1 min or

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Zeihl-Neelsen's carbol fuschin :-

Basic Fuschin (1gm) + Phenol (5gm) + Alcohol (95%)→10ml + distill water. →100 ml

wash

repert this till pink/red colour stop coming

Grading of smers (RNTCP) :

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No. of AFB seen in oil immersion fields Report

0/300 field NO AFB

1-2/300 field doutful, repert

1-9/ 100 field +1

1-9/10 field 2+

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1-9/1 Field 3+

10-more / 1 Field 4+

2% methylene blue 1 min, or malachite green

wash

drey → oil immersion → 100x

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Dry method Slide

Negatiu staining

Principle :- Indian Ink / negrosin of acidic)

a drop india Ink / negrosine

chromophore (-ve ion) H+ ion

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responsible for staining → stain only backgrow bez cell wall of bacteria is also - ve Charge so they repeal chromo

1 loop colony

mix

eg. Streptococcus pneumoniae → they seen bright

Crypto coccus neoformans /unstained against dark background.

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Spread with another slide (like PB)

H. influenzae

Klebsiella

Syphillis

dry

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oil immersies

100X

spore staimny

Heat fined slide

spore stain green and bacillus stain red

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spore→ Central oval Non buldging

Spherical

Terminal oval → Cl. tertium Buldging

spherical → Cl. tetani

sub terminal

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malachite green (5%)

5-6 min. heat till steam arises eg: Central → Bacillus anthraus

Sub terminal → Cl. perfringes

wash

(normal peptido-glycan) →Keratin

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Spore wall → cortex

Cell mem, (thick peptido-glycan)

Core

DNA

Ribosome

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glycolytic enzym

(0.5%) basic fuschin for 30sec.

wash drey

oil immersion spore represent a dormant resting stage in growth cycle of organiom.

100X

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Nutrient Broth (Pepton water + Y's mert extract)

+0.5% (glucose)

2-3% agar

glucose Broth

Nutrient agar

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Trichophyton mentagrophytes :- Clusters of microconidia macroconidia & cigar shaped with terminal rat tail filaments

Aspergillus fumigatus :- Cori vesicle is conical shaped Pristati Phialides arranged in Single rouw Conidia arises from upper 1324 Vesiclen

Aspergalle flas whole centia vesicle

Black Aspeye niger Condia

strep pneumoniae strep. pyogens Steph. epidermidis

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strep. viridans step. secprophyticus

Penicillin I stage branche

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