Immunofluorescence

1
Immunofluorescence

• Sayeed Ismail Khatib
• Lecturer
• Microbiology Immunology
• Ibn Sina National College for Medical Studies

2
Introduction

• Immunofluorescence is the labeling of antibodies
  or antigens with fluorescent dyes.
• This technique is sometimes used to make viral
  plaques more readily visible to the human eye.
• Immunofluorescent labeled tissue sections are
  studied using a fluorescence microscope.
• Fluorescein is a dye which emits greenish
  fluorescence under UV light. It can be tagged to
  immunoglobulin molecules.

3

• There are two ways of doing IF staining
• Direct immunofluorescence
• Indirect immunofluorescence
• 1. Direct immunofluorescence
• Ag is fixed on the slide
• Fluorescein labeled Abs are layered over it
• Slide is washed to remove unattached Abs
• Examined under UV light in an fluorescent
  microscope
• The site where the Ab attaches to its specific Ag
  will show apple green fluorescence
• Use Direct detection of Pathogens or their Ags
  in tissues or in pathological samples

4
Direct immunofluorescence
5

• Indirect immunofluorescence
• Indirect test is a double-layer technique
• The unlabelled antibody is applied directly to
  the tissue substrate
• Treated with a fluorochrome-conjugated
  anti-immunoglobulin serum

6

• Advantage over direct IF
• Because several fluorescent anti-immunoglobulins
  can bind to each antibody present in the first
  layer, the fluorescence is brighter than the
  direct test.
• It is also more time-efficient since it is only
  one signal labelled reagent, the
  anti-immunoglobulin, is prepared during the
  lengthy conjugation process

7
(No Transcript)
8
Indirect immunofluorescence of iron-regulated
cell wall mannoprotein FIT1 of S. cerevisiae
9
(No Transcript)
10
Confocal image to detect phosphorylated AKT
(green) in cardiomyocytes infected with
adenovirus
11
 Immunofluorescence image of Cryptosporidium
parvum oocysts       
12
                                                                                                                                                       
13
(No Transcript)
14
                                                
                                                 
Concentrated groundwater methanotrophic bacteria
on 0.2 m m filter labeled with fluorescent
antibodies.