Mouse CD4+/Granzyme B ELISpot Kit

PRODUCT SUMMARY
ELISpot kits are highly sensitive, microplate-based assays for the detection of cytokine secreting cells. This kit is designed for the detection and enumeration of mouse Granzyme B. Complete ELISpot kits are ready-to-run and require no assay development or refinement.

This ELISpot assay employs a capture antibody specific for mouse Granzyme B, pre-coated onto a PVDF-backed microplate. Appropriately stimulated cells are pipetted directly into the wells and the immobilized antibody in the immediate vicinity of the secreting cells binds secreted mouse Granzyme B. Following wash steps and incubation with a biotinylated detection antibody, alkaline-phosphatase conjugated to streptavidin is added. Unbound enzyme is subsequently removed by washing and a substrate solution (BCIP/NBT) is added. A blue-black colored precipitate forms at the sites of cytokine localization and appears as spots, with each individual spot representing an individual mouse Granzyme B secreting cell. The spots can be counted with an automated ELISpot reader system or manually using a stereomicroscope.

PRODUCT FEATURES

• Detect and quantitate individual cells secreting mouse Granzyme B
• High sensitivity - ELISpot assays can measure responses with frequencies well below 1 in 100,000 cells
• No in vitro expansion of cells required
• High-throughput - ELISpot assays use only a small number of primary cells

KIT CONTENTS

• Mouse Granzyme B Microplate
• Biotinylated Detection Antibody
• Streptavidin conjugated to Alkaline Phosphatase
• Dilution Buffers
• Wash Buffer Concentrate
• BCIP/NBT Chromogen
• Mouse Granzyme B Positive Control

OTHER REAGENTS REQUIRED

• Pipettes and pipette tips
• Deionized or distilled water
• Squirt bottle, manifold dispenser, or automated microplate washer
• 500 mL graduated cylinder
• 37 °C CO₂ incubator
• Sterile culture media
• Dissection microscope or an automated ELISpot reader