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  Fluorochromes Used In Immunology

Fluorochromes Used In Immunology:

1. Fluorescein isothiocyanate (i.e., FITC)

2. Lysamine-rhodamine B-200 Sulphonyl chloride

3. Rhodamine B isothiocyanate (i.e., RBITC)

Fluorochrome Labeling of Immunoglobulins:

Common considerations: Serum or gamma globulins can be employed for coupling.

1. Gamma globulin is superior since albumin in sera bind fast and

2. NH4SO4 is employed for precipitation of gamma globulin. It should be eliminated. Or else it competes with gamma globulin for dye.

3. The other factor is the weight ratio among fluorochrome & proteins.

4. pH of the atmosphere is significant. Alkaline pH improves the speed & quantity of coupling.

Removal Of Excess Labeled:

Dialysis, adsorption to active charcoal, precipitation, sephadex gel filtration G25 and ion exchange columns are employed.

 

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