Revvity, 64PARP1TPEH, HTRF Human and Mouse Total PARP1 Detection Kit, 10,000 assay points

Overview
PARP1 (Poly(ADP-ribose) polymerase 1) is a nuclear enzyme that plays a key role in DNA repair by utilizing NAD⁺ to synthesize poly(ADP-ribose) (PAR). Its overexpression in various cancers contributes to tumor progression. In cells with BRCA1 or BRCA2 deficiencies, PARP1 inhibition leads to apoptosis, making it a promising therapeutic target in cancer treatment.
HTRF assays offer many advantages over other technologies:

Homogeneous add-and-read format
No wash steps
Low background
Straightforward miniaturization from 96- or 384-well microplates to high density assay formats such as 384-well low volume and 1536-well plates
Stable signal, providing flexibility in time of readout or size of assays
How it works
Total PARP1 assay principle
The Total PARP1 assay quantifies the expression level of PARP1 in a cell lysate. Unlike Western Blot, the assay is entirely plate-based and does not require gels, electrophoresis, or transfer. The 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 PARP1 this enables an immune-complex formation involving both labeled antibodies, and which brings the donor fluorophore into close proximity to the acceptor, thereby generating a FRET signal.
Its intensity is directly proportional to the concentration of total protein present in the sample, and provides a means of assessing the protein's phosphorylation state under a no-wash assay format.

 

Total PARP1 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 PARP1 detection reagents. This protocol allows for the cells' viability and confluence to be monitored.

Total PARP1 one-plate assay protocol
Detection of Total PARP1 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 4
Molecular Modification-Total
Product Group-Kit
Sample Volume-16 µL
Shipping Conditions-Shipped in Dry Ice
Target-PARP1
Target Class-Phosphoproteins
Target Species-Human Mouse
Technology-TR-FRET
Therapeutic Area-Oncology
Unit Size-10,000 assay points