Revvity, 64HSP70TPEH, HTRF Human and Mouse Total HSP70-1A/1B Detection Kit, 10,000 assay points

Overview
HSP70-1A and HSP70-1B, also known as HSPA1A and HSPA1B, are the major stress-inducible 70-kDa heat shock proteins. These highly conserved molecular chaperones play a critical role in maintaining protein quality control and cellular homeostasis under stressful conditions such as heat shock, oxidative stress, exposure to heavy metals, or proteotoxic drugs.
HSP70-1A/1B are also potent anti-apoptotic proteins, making them essential for the growth and survival of cancer cells. Elevated expression levels of these proteins are observed in various cancer types and are associated with poor prognosis, recurrence, and resistance to treatment. Consequently, targeting HSP70-1A/1B represents a promising strategy to sensitize tumors to therapy.
Conversely, HSP70-1A/1B have been shown to protect the brain from a range of nervous system injuries by inhibiting both cell death and immune responses. As a result, pharmaceutical compounds capable of inducing HSP70 expression are being explored as potential treatments for neurological diseases.
How it works
Total HSP70-1A/1B assay principle
The Total HSP70-1A/1B assay measures the levels of HSP70-1A/1B in cells. Unlike Western Blot, this assay is entirely plate-based and does not require gels, electrophoresis, or protein transfer. It utilizes two antibodies: one labeled with a donor fluorophore and the other with an acceptor. Both antibodies are highly specific to distinct epitopes on the target protein.
In the presence of HSP70-1A/1B, an immune complex forms involving both labeled antibodies, bringing the donor fluorophore into close proximity with the acceptor. This interaction generates a FRET signal, the intensity of which is directly proportional to the concentration of total protein in the sample. The assay also enables assessment of the protein’s phosphorylation state, all within a no-wash assay format.

 

Total HSP70-1A/1B 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 Total HSP70-1A/1B HTRF detection reagents. This protocol allows for the cells' viability and confluence to be monitored.

Total HSP70-1A/1B one-plate assay protocol
Detection of Total HSP70-1A/1B 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 allows miniaturization while maintaining robust HTRF quality. Application-Cell Signaling
Brand-HTRF
Detection Modality-HTRF
Lysis Buffer Compatibility-Lysis Buffer 1
Molecular Modification-Total
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target-HSP70 1A/1B
Target Class-Phosphoproteins
Target Species-Human Mouse
Technology-TR-FRET
Therapeutic Area-Central Nervous System Oncology
Unit Size-10,000 assay points