By

ARCHANA NAGAR
&
GIRISH TANK
1. Quality assurance
2. Quality control
3. Quality control of AFB staining
4. Quality control of Gram staining



Quality assurance:
It is a system which assure that the
overall activities in the laboratory are
being performed as designed by
particular standards.
Quality control:
Quality control deals with the
system which accepts or rejects
any activities or parameter
which affects the quality of
product and thus prevent
quality deficiency.
A control slide should be included with each run of
stain this will verify the correct performance of the
procedure as well as the staining intensity of the
acid fast organisms.
SLIDE PREPARATION:- QC slides may be
prepared as follows-
1. Prepare seprate suspension approximately equal to a no .
1 Mc farland turbidity standard of
E.coli ATCC 25922(negative control) and
mycobacterium tuberculosis ATCC H37RU (positive
control) in approximately 1.0 ml of physiological saline
or sterile water in a biological safety cabinet.
McFarland Standards
McFarland turbidity standards are
used to standardize the approximate
number of bacteria in a liquid
suspension by visually comparing the
turbidity of a test suspen-sion with the
turbidity of a McFarland standard.
2. LABLING:-lable control slides with name(eg.
AFB control,like positive and negative
control).Record date of preparation and
preparers name appropriately.
3. Using a insulin syring or loop place a drop of the
negaive control suspension near the end of the
glass slide.
4. Repeat step 3 with the suspension of the positive
control ,but place the drop half way between
positive negative control and the slide lable.
5. Allow smears to air dry in the biological safety
cabinet.
6. Heat fix the smears.
7. Store control slides in a labeled slide box.

Staining Procedure:-
8.Flood slide with Carbol Fuchsin
9.Hold a flame beneath the slide until steam
appears but do not allow it to boil
10.Allow hot slide to sit for 3 to 5 minutes, rinse
with tap water
11.Flood slide with acid fast decolorizer
12.Allow to sit 1 minute, rinse with tap water
13.Flood slide with Methylene Blue
14.Allow to sit 1 minute, rinse with tap water
15.Blot dry
16.View under oil immersion lens

Acid Fast Staining
EVALUATION:-

1. Control slide should be reviewed before
the patient smears are read to confirm
that the mycobacteria stain acid fast.
a. Carbolfuchsin
i. mycobacterium tuberculosis (ATCC
H37RU ): red or magenta against a blue
background
ii. E.coli (ATCC 25922): blue.
b.carbolfuchsin
i. positive control is not stained red
ii.negative control remains red after
decolorization
iii.background is not properly decolorized.
C. When the problem is resolved,re-
stain a control slide as well as all
patients slides from the problem
run.
d. Record the QC slide results on the
appropriate log sheet.Fill out the
cosecutive action if there are any of
the above mentioned problem.
QUALITY CONTROL OF
GRAM STAINING
1. Prepare seprate suspension approximately
equal to a no . 1 Mc farland turbidity
standard of
SLIDE PREPARATION:- QC slides may be
prepared as follows-
Positive control: Staphylococcus aureus
(ATCC 25923)

Negative control: Escherichia coli(ATCC 25922)

in approximately 1.0 ml of physiological saline or sterile
water in a biological safety cabinet.


2. LABLING:-lable control slides with name(eg.
GRAM control,like positive and negative
control).Record date of preparation and
preparers name appropriately.
3. Using a insulin syring or loop place a drop of
the negaive control suspension near the end
of the glass slide.
4. Repeat step 3 with the suspension of the
positive control ,but place the drop half way
between positive negative control and the
slide lable.

5. Allow smears to air dry in the biological safety
cabinet.
6.Heat fix the smears.
7. store control slides in a labeled slide box.
Staining Procedure:-
8.On a rack, flood with filtered crystal violet
10 sec
9. Wash briefly in water to remove excess crystal
violet
10. Flood with Grams iodine 10 sec
11. Wash briefly in water, do not let the section
dry out.
12.Decolourise with acetone until the moving
dye front has passed the lower edge of the
section
13. Wash immediately in tap water
14. Counterstain with safranin 15 seconds






QUALITY CONTROL:-
A. Check appearance of reagents daily
If crystal violet has precipitate or crystal sediment ,
refilter before use .
B. Daily and when a new lot is used , prepare a smear of
E.coli (ATCC 25922) and Staphylococcus aureus
(ATCC 25923)
E.coli (GNB - Pink color)
Staphylococcus aureus (GPC - deep violet color)
Dr.T.V.Rao MD 21
Dr.T.V.Rao MD 22
Good Quality Poor Quality
THANK YOU