DESCRIPTION

PARK7, also named as DJ1, belongs to the peptidase C56 family. It protects cells against oxidative stress and cell death. PARK7 plays a role in regulating expression or stability of the mitochondrial uncoupling proteins SLC25A14 and SLC25A27 in dopaminergic neurons of the substantia nigra pars compacta and attenuates the oxidative stress induced by calcium entry into the neurons via L-type channels during pacemaking. It eliminates hydrogen peroxide and protects cells against hydrogen peroxide-induced cell death. PARK7 has cell-growth promoting activity and transforming activity. It may function as a redoxsensitive chaperone. It's precursor undergoes a cleavage of a C-terminal peptide and subsequent activation of protease activity in response to oxidative stress.

PRINCIPLE OF THE ASSAY

An anti-tag antibody is pre-coated onto the bottom of wells. After adding antigen or samples, Tagged-Capture antibody and HRP-conjugated detection antibody, a sandwich complex is formed in the solution. TMB acts as a HRP substrate, and the solution color will change from colorless to blue. A stop solution containing sulfuric acid turns the 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	4.91	0.08	1.63
2	8	1.28	0.02	1.56
3	8	0.67	0.02	2.99

Inter-assay Precision

Sample	n	Mean (ng/mL)	SD	CV%
1	16	4.98	0.10	2.01
2	16	1.30	0.03	2.31
3	16	0.69	0.06	8.70

RECOVERY

The recovery of human PARK7/DJ-1 spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type		Average% of Expected	Range (%)
Human plasma	1:16	121	117-125
	1:32	115	105-123
Cell lysate	1:6,400	96	82-104
	1:12,800	99	94-103
Human cerebrospinal fluid (CSF)	1:8	99	95-102
	1:16	82	73-96

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 human plasma was initially diluted 1:2. The cell lysate was initially diluted 1:800. The human cerebrospinal fluid was initially diluted 1:4.)

		Human plasma (SampleDiluent PT 3-ef)	Cell lysate (SampleDiluent PT 4B1)	Human cerebrospinal fluid(Sample Diluent PT 4B1)
1:2	Average% of Expected	100	100	100
	Range (%)	-	-	-
1:4	Average% of Expected	104	103	108
	Range (%)	103-105	102-104	102-115
1:8	Average% of Expected	112	109	117
	Range (%)	100-123	99-119	110-124
1:16	Average% of Expected	105	106	106
	Range (%)	100-110	93-119	97-116

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human PARK7/DJ-1 One-Step ELISA Kit produced at Sunlong Medical™.