DESCRIPTION

CLPX is a kind of protein that plays an important role in mitochondria. It belongs to AAA+ protein family and mainly participates in the degradation process of protein. As a chaperone subunit, CLPX and CLPP protease subunit together form a clxp protease complex, which relies on ATP to recognize, expand and degrade specific target proteins. CLPX is expressed in many tissues of human body, such as liver and testis. It plays a key role in maintaining intracellular protein homeostasis, stress response and mitochondrial function.

PRINCIPLE OF THE ASSAY

Sandwich ELISA structure (Detection antibody labeled with HRP) A capture antibody is pre-coated onto the bottom of wells which binds to analyte of interest. A detection antibody labeled with HRP also binds to the analyte. TMB acts as the HRP substrate and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns solution yellow. The color intensity is proportional to the quantity of bound protein which is measurable at 450 nm with the correction wavelength set at 630 nm.

PRECISION

Intra-assay Precision (Precision within an assay) Three samples of known concentration were tested 8 times on one plate to assess intra-assay precision. Inter-assay Precision (Precision between assays) Three samples of known concentration were tested in 16 separate assays to assess inter-assay precision.

Intra-assay Precision

Sample	n	Mean (ng/mL)	SD	CV%
1	8	21.70	1.83	8.41
2	8	5.08	0.35	6.91
3	8	2.70	0.10	3.88

Inter-assay Precision

Sample	n	Mean (ng/mL)	SD	CV%
1	16	20.79	1.98	9.55
2	16	5.16	0.31	5.92
3	16	2.80	0.14	4.92

RECOVERY

The recovery of human CLPX spiked to three different levels throughout the range of the assay in cell lysate was evaluated.

Sample Type		Average% of Expected	Range (%)
Cell lysate	1:128	86	83-88

LINEARITY

To assess the linearity of the assay, samples were diluted with the appropriate Sample Diluent to produce samples with values within the dynamic range of the assay. (The cell lysate was initially diluted 1:16.)

		Cell lysate
1:2	Average% of Expected	100
	Range (%)	-
1:4	Average% of Expected	102
	Range (%)	95-118
1:8	Average% of Expected	99
	Range (%)	75-127
1:16	Average% of Expected	96
	Range (%)	70-129

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human CLPX Sandwich ELISA Kit produced at Sunlong Medical™.