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Koronavírus diagnosztika
Flow citometria
Klinikai kémia
Transzplantációs immungenetika
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1031 Budapest, Szentendrei út 176.
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Műszerek és eszközök
Ki-67 + pHH3
Gyártó: Biocare Medical

Catalog Number: API 3198DS AA
Description: 6.0 ml, prediluted
Dilution: Ready-touse
Diluent: N/A

Intended Use:
For In Vitro Diagnostic Use
Ki-67 + pHH3 is a cocktail of mouse and rabbit monoclonal antibodies that is intended for laboratory use in the qualitative identification of Ki-67 and phosphohistone H3 (Ser10) proteins by immunohistochemistry (IHC) in formalin-fixed paraffin-embedded (FFPE) human tissues. The clinical interpretation of any staining or its absence should be complemented by morphological studies using proper controls and should be evaluated within the context of the patient’s clinical history and other diagnostic tests by a qualified pathologist.
Summary and Explanation:
The Ki-67 nuclear antigen is associated with cell proliferation and found throughout the cell cycle; though not in G0 phase (1,2). The assessment of Ki-67 proliferation in breast cancers has shown the Ki-67 labelling index is an important predictor of survival (3).
Microscopic evaluation of mitotic figures on H&E is a routine procedure in the assessment of the tumor grades (4). However, the counting of mitosis is manual and time consuming with assorted difficulties as well as variabilities between interobserver assessments (5). Histone H3 phosphorylation at Serine10 (pHH3) is in association with mitotic chromatin condensation in late G2 and M phase of the cell cycle. pHH3 can distinguish mitosis from apoptotic nuclei (6). The immunohistochemical staining of Serine10-pHH3 has been reported to be comparable to mitotic figures in the H&E section (7-10).
Immunohistochemical studies have shown immunostaining using anti- Ki-67 and anti-pHH3 antibodies provided improved prediction of recurrence of tumors and may become effective ancillary tools in deciding on optimal therapeutic management (11). Other studies have shown Ki-67 and pHH3 can be useful in separating malignant melanoma from benign nevi (12).
Principle of Procedure:
This product is a primary antibody cocktail of mouse and rabbit antibodies, which may be used in a Multiplex IHC staining procedure to produce a two-color stain. Following application of the primary antibody cocktail to the tissue
sample, detection is performed by separate secondary antibodies specific for each species (i.e. mouse or rabbit)
of the primary antibody cocktail, which are conjugated to horseradish peroxidase (HRP) or alkaline phosphatase (AP) enzymes. Visualization is accomplished by the application of chromogenic substrates (DAB and Warp Red), which are enzymatically activated (by HRP or AP, respectively) to produce a colored reaction product at the antigen site. The specimen may be counterstained and coverslipped. Results are interpreted using a light microscope.

Reagent Provided:
Ki-67 + pHH3 is provided as a prediluted antibody cocktail of anti- Ki-67 and anti-pHH3 antibodies in buffer with carrier protein and preservative.

Antibody anti-Ki-67 anti-pHH3
Clone MIB-1 BC37
Source Mouse monoclonal Rabbit monoclonal
Isotype IgG/kappa IgG
Epitope/ Antigen 1002 bp Ki-67 cDNA fragment PhosphoSer10 of Histone H3
Cellular Localization Nuclear Nuclear (mitotic figure)
Staining Red
(Warp Red) Black (Deep Space)

Storage and Stability:
Store at 2°C to 8°C. Do not use after expiration date printed on vial. If reagents are stored under conditions other than those specified in the package insert, they must be verified by the user. Diluted reagents should be used promptly; any remaining reagent should be stored at 2°C to 8°C.
Known Applications:
Immunohistochemistry (formalin-fixed paraffin-embedded tissues)
Species Reactivity: Human; others not tested
Positive Tissue Control: Melanoma or colon cancer
Protocol Recommendations:
Peroxide Block:
Block for 5 minutes with Biocare's Peroxidazed 1.
Pretreatment: Perform heat retrieval using Biocare’s Diva Decloaker. Refer to the Diva Decloaker data sheet for specific instructions.
Protein Block: Incubate for 10 minutes at RT with Biocare's Background Punisher.
Primary Antibody: Incubate for 30 minutes at RT.
Double Stain Detection: Incubate for 30 minutes at RT using Biocare's MACH 2 Double Stain 1.
Chromogen (1): Incubate for 5 minutes at RT with Biocare's Deep Space Black.
Chromogen (2): Incubate for 5-7 minutes at RT with Biocare's Warp Red. Rinse in deionized water.
Counterstain with hematoxylin. Rinse with deionized water. Apply Tacha's Bluing Solution for 1 minute. Rinse with deionized water.

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