Unit 5: Enzymes, Substrates, and Chromogens

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2 common flurochromes used in IHC

Fluorescein isothiocynate (FITC), rhodamine

2 common enzymes used in IHC

horseradish peroxidase, alkaline phosphatase

Substrate in horseradish peroxidase reaction

hydrogen peroxide

How can false positive reactions with endogenous peroxidase interfering with horseradish peroxidase be blocked?

hydrogen peroxide

Substrate of alkaline phosphatase reaction

naphthol phosphate ester

How is chromogen made?

colorless reagent that is converted to a stable-colored end product

T/F chromogens are electron donors

true

T/F chromogens are electron acceptors

false

Chromogens used most frequently with HRP are..

DAB, AEC

What is DAB oxidized by?

hydrogen peroxide

WHat is a chromogen?

a stable-colored end product

What makes the site of the antigen/antibody complex?

chromogen

Metals that can increase intensity of the DAB reaction

nickel, cobalt

T/F upon oxidation AEC forms a red end product

true

T/F upon oxidation AEC forms a brown end product

false

T/F AEC fades over time

true

What color end product is 4-chloro-1-naphthol (CN)?

blue

Which reaction tends to diffuse from site of precipitation

4-chloro-1-naphthol (CN)

What color end product is P-phenylenediamine dihydrochloride (Hanker-Yates reagent)?

blue-black

T/F phosphate buffered saline (PBS) can be used as a rinsing buffer when using alkaline phosphate

false

Recommended rinsing buffer for alkaline phosphate system

TRIS

T/F you should use DAB for melanoma

false

What color betagalactosidase?

turquoise

What color is glucose oxidase reaction?

blue