Human Xeroderma Pigmentosum, Complementation Group D (XPD) ELISA Kit

Principle of the Assay

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

ATP-dependent 5'-3' DNA helicase, component of the general transcription and DNA repair factor IIH (TFIIH) core complex, which is involved in general and transcription-coupled nucleotide excision repair (NER) of damaged DNA and, when complexed to CAK, in RNA transcription by RNA polymerase II. In NER, TFIIH acts by opening DNA around the lesion to allow the excision of the damaged oligonucleotide and its replacement by a new DNA fragment. The ATP-dependent helicase activity of XPD/ERCC2 is required for DNA opening. In transcription, TFIIH has an essential role in transcription initiation. When the pre-initiation complex (PIC) has been established, TFIIH is required for promoter opening and promoter escape. Phosphorylation of the C-terminal tail (CTD) of the largest subunit of RNA polymerase II by the kinase module CAK controls the initiation of transcription. XPD/ERCC2 acts by forming a bridge between CAK and the core-TFIIH complex. Involved in the regulation of vitamin-D receptor activity. As part of the mitotic spindle-associated MMXD complex it plays a role in chromosome segregation. Might have a role in aging process and could play a causative role in the generation of skin cancers.

GENE ID	2068
SWISS PROT	P18074
SYNONYMS	CXPD; BTF2 p80; EM9; TTD; ERCC2; TFIIH basal transcription factor complex 80 kDa; De Sanctis-Cacchione; Excision Repair Cross-Complementing Rodent Repair Deficiency 2

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 XPD were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level XPD 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 0.056ng/mL.
ASSAY RANGE	0.156-10ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of XPD. No significant cross-reactivity or interference between XPD and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between XPD and all analogs, therefore, cross reactivity may still exist.