Revvity, 64HPK1TPEG, HTRF Human Total HPK1 Detection Kit, 500 Assay Points

Overview
Hematopoietic progenitor kinase 1 (HPK1) is a serine/threonine kinase primarily involved in the regulation of immune cell signaling. It plays a crucial role in T-cell receptor (TCR) signaling by modulating downstream pathways that influence T-cell activation, proliferation, and differentiation. HPK1 negatively regulates TCR signaling by phosphorylating adaptor proteins such as SLP-76, which dampens T-cell responses. Additionally, HPK1 is involved in B-cell receptor (BCR) signaling, impacting B-cell activation and function. It also contributes to the innate immune response by modulating pathways in myeloid cells, such as dendritic cells and macrophages, affecting cytokine production and cell migration. Beyond immune cells, HPK1 has been implicated in the regulation of apoptosis and cell cycle progression, influencing hematopoietic cell homeostasis. Due to its regulatory roles, HPK1 is a potential therapeutic target for immune-related disorders and cancers.
How it works
Total HPK1 assay principle
The Total HPK1 assay quantifies the expression level of HPK1 in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The Total HPK1 assay uses two labeled antibodies, one coupled to a donor fluorophore and the other to an acceptor. Both antibodies are highly specific for a distinct epitope on the protein. In the presence of HPK1 in a cell extract, the addition of these conjugates brings the donor fluorophore into close proximity with the acceptor and thereby generates a FRET signal. Its intensity is directly proportional to the concentration of the protein present in the sample and provides a means of assessing the protein's expression under a no-wash assay format.

 

Total HPK1 two-plate assay protocol
The two-plate protocol involves culturing cells in a 96-well plate before lysis, then transferring lysates into a 384-well low volume detection plate before the addition of HTRF Total HPK1 detection reagents. This protocol allows for the cells' viability and confluence to be monitored.

Total HPK1 one-plate assay protocol
Detection of Total HPK1 with HTRF reagents can be performed in a single plate used for culturing, stimulation, and lysis. No washing steps are required. This HTS designed protocol facilitates miniaturization while maintaining robust HTRF quality. Application-Cell Signaling
Brand-HTRF
Buffer/Solvent-Lysis Buffer 4
Detection Modality-HTRF
Host Species-Human
Molecular Modification-Total
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target-HPK1
Target Class-Phosphoproteins
Technology-TR-FRET
Therapeutic Area-Inflammation Oncology
Unit Size-500 assay points