DESCRIPTION

Phosphoglycerate kinase 1 (PGK1) is a key glycolytic enzyme encoded by an X-linked gene (Xq13), ubiquitously expressed across human cells. It catalyzes the reversible transfer of a phosphate group from 1,3-bisphosphoglycerate to ADP, generating 3phosphoglycerate and ATP-the first ATP-producing step of glycolysis. Structurally, PGK1 is a ~45 kDa monomeric protein with two Rossmann-fold domains connected by a hinge; a hinge-bending motion brings substrates into a reactive closed conformation. Beyond canonical metabolism, PGK1 exhibits moonlighting functions, including autophagy initiation, DNA repair, and angiogenesis regulation. Its role in cancer is context-dependent: elevated intracellular PGK1 promotes tumor proliferation, the Warburg effect, and chemoradiotherapy resistance, whereas extracellular PGK1 can suppress malignancy by inhibiting angiogenesis. PGK1 abnormalities are also linked to hemolytic anemia, myopathy, and neurological disorders.

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	5.44	0.08	1.47
2	8	1.37	0.01	0.73
3	8	0.69	0.01	1.45

Inter-assay Precision

Sample	n	Mean (ng/mL)	SD	CV%
1	16	5.45	0.07	1.28
2	16	1.37	0.02	1.46
3	16	0.69	0.01	1.45

RECOVERY

The recovery of human PGK1 spiked to three different levels throughout the range of the assay in various matrices was evaluated.

Sample Type		Average% of Expected	Range (%)
Human serum	1:2	86	83-90
Cell lysate	1:800	96	94-98
	1:1,600	97	94-98

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 1:100.)

		Human serum(Sample Diluent PT 1)	Cell lysate(Sample Diluent PT 4B1)
1:2	Average% of Expected	100	100
	Range (%)	-	-
1:4	Average% of Expected	112	91
	Range (%)	103-122	91-92
1:8	Average% of Expected	121	85
	Range (%)	116-126	85-86
1:16	Average% of Expected	117	82
	Range (%)	105-129	82-83

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human PGK1 produced at Sunlong Medical™.

Ordering & shipping

• Minimum order: 1 kit
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