DESCRIPTION

Human meningioma-expressed antigen 5 (MGEA5) has two putative domains including protein O-GlcNAcase domain and histone acetyltransferase domain, therefore it is often called bifunctional protein NCOAT. Three isoforms of MEGE5 are produced by alternative splicing. MGEA5 was found to be regulated to reduce the state of glycosylation of transcitpional activators while increasing the acetylation of histones to allow for the concerted activation of eukaryotic gene transcription, for instance, acetylation of Lys8 of histone H4 and Lys 14 of histone H3 are resulted from acetyltransferase activity. In addition, single nucleotide polymorphism in MGE5A is associated with type 2 diabetes in Mexican Americans.

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	7.66	0.43	5.64
2	8	1.92	0.06	2.95
3	8	0.98	0.07	7.63

Inter-assay Precision

Sample	n	Mean (ng/mL)	SD	CV%
1	16	7.69	0.39	5.12
2	16	1.96	0.10	5.03
3	16	0.98	0.07	7.37

RECOVERY

The recovery of human MGEA5 spiked to three different levels throughout the range of the assay in cell lysate was evaluated.

Sample Type		Average% of Expected	Range (%)
Cell lysate	1:64	81	81-82

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 1:4.)

		Cell lysate
1:2	Average% of Expected	100
	Range (%)	-
1:4	Average% of Expected	84
	Range (%)	83-85
1:8	Average% of Expected	79
	Range (%)	74-84
1:16	Average% of Expected	96
	Range (%)	86-105

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human Human MGEA5 Sandwich ELISA Kit produced at Sunlong Medical™.