Dako Autostainer User Manual download pdf (Page 10)

Do not print slide labels while the Autostainer is running.

Immunohistochemistry is a multi-step diagnostic process that requires specialized

training in the selection of the appropriate reagents, tissue selection, fixation, and

processing, preparation of the IHC slide, and interpretation of the staining results.

Tissue staining is dependent on the proper handling and processing of tissues prior to

staining. Improper fixation, freezing, thawing, washing, drying, heating, sectioning or

contamination with other tissues or fluids may produce artifacts, antibody trapping, or

false-negative results. Inconsistent results may be due to variations in fixation and

embedding methods, or to inherent irregularities within the tissue. Excessive or

incomplete counterstaining may compromise proper results.

Use of old or unbuffered fixatives, or exposure of tissues to excessive heat (greater than

60°C) during processing may result in decreased staining sensitivity.

Normal/non-immune sera from the same animal source as the secondary antisera used

in blocking steps may cause false-negative or false-positive results due to auto-

antibodies or natural antibodies. False-positive results may be seen due to non-

immunologic binding of reagents to tissue sections. In some case the application of an

alternate blocking reagent prior to incubation with the primary antibody may be useful for

reducing background. A recommended blocking reagent is Dako Protein Block Serum-

Free (Code No. X0909).

Unexpected negative reactions in poorly differentiated neoplasms may be due to loss or

marked decrease of antigen expression or nonsense mutation in the gene(s) coding for

the antigen. Unexpected positive staining in tumors may be from expression of an

antigen not usually expressed in morphologically similar normal cells, or from

persistence or acquisition of an antigen in a neoplasm that develops morphologic and

immunohistochemical features associated with another cell lineage (divergent

differentiation). Histopathologic classification of tumors is not an exact science and some

literature reports of unexpected staining may be controversial.

The clinical interpretation of any positive staining or its absence should be

complemented by morphological and histological studies with proper controls.

Evaluations should be made within the context of the patient’s clinical history and other

diagnostic tests. It is the responsibility of a qualified pathologist who is familiar with the

antibodies, reagents and methods used to interpret the stained preparation. Staining

must be performed in a certified licensed laboratory under the supervision of a

pathologist who is responsible for reviewing the stained slides and assuring the

adequacy of positive and negative controls.

Reagents may demonstrate unexpected reactions in previously untested tissues. The

possibility of unexpected reactions in tested tissue groups cannot be completely

eliminated due to biological variability of antigen expression in neoplasms, or other

pathological tissues. Contact your local Dako representative with documented

unexpected reactions. Tissues from people infected with hepatitis B virus and containing

hepatitis B surface antigen (HBsAg) may exhibit nonspecific staining with horseradish

peroxidase.

2 Autostainer Handbook

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Dako Autostainer User Manual Page 10

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