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Western Blot analysis of Perforin (mouse) mAb (CB5.4) (Prod. No. ALX-804-057): Lane 1: MW Marker, Lane 2: CTLL-2 cell lysate, Lane 3: 3T3 cell lysate. Western Blot analysis of Perforin 1 expression in CTLL-2 cell lysate and absence of perforin in the mouse fibroblast cell line 3T3.
Figure: Detection of perforin in an individual mouse T cell. Perforin is located in cytoplasmic granules.
Method: A mouse T cell clone was plated onto polylysine treated glass slides, fixed and permeabilized in methanol at -20oC for 5 min, then in acetone at -20oC for 30 sec. After 3 washings in PBS, 0.1% BSA, slides were incubated with 5 µg/ml of CB5.4 antibody, washed in PBS, incubated in anti-rat IgG-HRP, washed in PBS, and developed in appropriate substrate solution (DAB or AEC).
Schematic structure of mouse perforin (recombinant, aa. 98-534). The antibody recognizes an epitope within aa. 402-534.
Figure: Detection of perforin in the mouse T cell clone B6.1. (lane 1). Absence of perforin in the mouse fibroblast cell line NIH.3T3 (lane 2). The protein migrates as a 66 kDa species.
Method: Cell extracts from the T cell clone B6.1 (2×106) were resolved by SDS-PAGE under reducing conditions, transferred to nitrocellulose and probed with the CB5.4 antibody at 1 µg/ml. Proteins were visualized using a peroxidase-conjugated antibody to rat IgG and a chemiluminescence detection system.
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RUO – Research Use Only |
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