Mouse Caspase 1 (CASP1) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to CASP1. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to CASP1. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After the TMB substrate solution is added, only those wells that contain CASP1, biotin-conjugated antibody, and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution, and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of CASP1 in the samples is then determined by comparing the O.D. of the samples to the standard curve.

For Use with serum, plasma, and cell culture supernatants. For Research Use Only. Not for use in diagnostic procedures.

Target Information

Thiol protease involved in a variety of inflammatory processes by proteolytically cleaving other proteins, such as the precursors of the inflammatory cytokines interleukin-1 beta (IL1B) and interleukin 18 (IL18) as well as the pyroptosis inducer Gasdermin-D (GSDMD), into active mature peptides (PubMed:21147462, PubMed:32109412). Plays a key role in cell immunity as an inflammatory response initiator: once activated through formation of an inflammasome complex, it initiates a pro-inflammatory response through the cleavage of the two inflammatory cytokines IL1B and IL18, releasing the mature cytokines which are involved in a variety of inflammatory processes (PubMed:21147462). Cleaves a tetrapeptide after an Asp residue at position P1 (PubMed:21147462). Also initiates pyroptosis, a programmed lytic cell death pathway, through cleavage of GSDMD (PubMed:32109412). In contrast to cleavage of interleukins IL1B and IL1B, recognition and cleavage of GSDMD is not strictly dependent on the consensus cleavage site but depends on an exosite interface on CASP1 that recognizes and binds the Gasdermin-D, C-terminal (GSDMD-CT) part (PubMed:32109412). Upon inflammasome activation, during DNA virus infection but not RNA virus challenge, controls antiviral immunity through the cleavage of CGAS, rendering it inactive (PubMed:28314590). In apoptotic cells, cleaves SPHK2 which is released from cells and remains enzymatically active extracellularly (By similarity).

GENE ID	12362
SWISS PROT	P29452
SYNONYMS	ICE; IL1BC; IL1-BC; P45; Interleukin 1 Beta Converting Enzyme; Apoptosis-Related Cysteine Peptidase; Interleukin-1 beta convertase

Materials Supplied

Kit Components	96 Wells Quantity/Size
Pre-coated, ready-to-use 96-well strip plate	1 plate
Plate sealer for 96 wells	2
Standard	2 tubes
Diluent buffer	1 bottle
Detection Reagent A	1 bottle
Detection Reagent B	1 bottle
TMB Substrate	1 tube
Stop Solution	1 tube
Wash Buffer (30 ℅ concentrate)	1 tube
Product data sheet	1 copy

Storage

The TMB Substrate, Wash Buffer (30X concentrate), and the Stop Solution should be stored at 4°C upon receipt, while the other items should be stored at -20°C.

Performance Characteristics

REPEATABILITY	Intra-assay Precision (Precision within an assay): 3 samples with low, middle, and high-level CASP1 were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level CASP1 were tested on 3 different plates, with 8 replicates in each plate. CV(%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%
SENSITIVITY	The minimum detectable dose was 0.059ng/mL.
ASSAY RANGE	0.156-10ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of CASP1. No significant cross-reactivity or interference between CASP1 and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between CASP1 and all analogs, therefore, cross reactivity may still exist.