Human Lysine Specific Demethylase 4A (KDM4A) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to KDM4A. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to KDM4A. 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 KDM4A, 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 KDM4A 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

Histone demethylase that specifically demethylates 'Lys-9' and 'Lys-36' residues of histone H3, thereby playing a central role in histone code (PubMed:26741168). Does not demethylate histone H3 'Lys-4', H3 'Lys-27' nor H4 'Lys-20'. Demethylates trimethylated H3 'Lys-9' and H3 'Lys-36' residue, while it has no activity on mono- and dimethylated residues. Demethylation of Lys residue generates formaldehyde and succinate. Participates in transcriptional repression of ASCL2 and E2F-responsive promoters via the recruitment of histone deacetylases and NCOR1, respectively.

GENE ID	9682
SWISS PROT	O75164
SYNONYMS	JMJD2; JMJD2A; JHDM3A; JmjC domain-containing histone demethylation protein 3A; Jumonji domain-containing protein 2A

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 KDM4A were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level KDM4A 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 0.128ng/mL.
ASSAY RANGE	0.312-20ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of KDM4A. No significant cross-reactivity or interference between KDM4A and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between KDM4A and all analogs, therefore, cross reactivity may still exist.