This is only for study purposes.

1. IMMUNOFLUORESCENSE ASSAY
PRESENTER:
SAMIR MOIRANGTHEM
Msc. MLT, Medical Microbiology
RIPANS

2. IMMUNOFLUORESCENCE
• Fluorescence is the property of certain dyes which
absorb rays of one particular wavelength (
ultraviolet) and emits rays with a different
wavelength( visible light).
• It's is a method of determining the location of
antigen (or antibody) in a tissue section or smear
using a specific antibody (or antigen) labeled with
flurochrome.
• commonly used dyes are fluorescin isothiocyanate ,
lissamine rhodamine.

3. HISTORY
• First described in 1942 and refined by Albert Coons
in 1950.
• Coons and his colleagues showed that fluorescent
dyes can be conjugated to antibodies and these
"labelled" antibodies can be used to detect antigen
in tissues.

5. TYPES
• 1. DIRECT IMMUNOFLUORESCENSE TEST
• 2. INDIRECT IMMUNOFLUORESCENSE TEST

6. • 1. DIRECT IMMUNOFLUORESCENCE TEST
• PRINCIPLE:
• The specific antibodies tagged with fluorescent
dye(i.e.labelled antibodies) are used for detection
of unknown antigen in a specimen.
• If antigen is present, it reacts with labelled
antibodies and fluorescence can be observed
under ultraviolet light of fluorescent microscope.

9. USES

10. DISADVANTAGE
• SEPERATE SPECIFIC FLUORESCENT LABELLED
ANTIBODY HAS TO BE PREPARED AGAINST EACH
ANTIGEN TO BE TESTED.

11. 2. INDIRECT IMMUNOFLUORESCENCE
TEST
• used for detection of antibodies in serum or other body
fluids.
• PRINCIPLE : A known antigen is fixed on a slide. The
unknown antibody(serum) is applied to the slide.
• if antibody (globulin) is present in the serum, it attaches to
known antigen on the slide.
• for detection of this antigen-antibody reaction ,
fluorescein-tagged antibody to human globulin is added.
• in positive test, fluorescence occurs under ultraviolet light.

13. ADVANTAGES
• A single antihuman globulin fluorescent
conjugate can be employed for detection of
antibody to any antigen.
• All antibodies are globulin in nature,
therefore, antihuman globulin attaches to
all antibodies.
• This has overcome the disadvantage of
direct immunofluorescence test.

15. Sandwich' Technique of Immunofluorescence
Antigen being in the middle with labelled and unlabelled antibody
on either side, forms a sandwich. This is used for detection of
antibodies.
Antibody
+
Antigen
+
Labelled antibody

17. REFERENCE
Textbook Of Microbiology by C.P. Baveja
3rd edition

18. THANK YOU