DESCRIPTION

Used for the quantitative determination of Human Anti-Monkeypox virus IgG (A29L) concentration in serum and plasma.

PRINCIPLE OF THE ASSAY

This assay employs the quantitative competitive enzyme immunoassay technique. Recombinant Monkeypox virus A29L antigen has been pre-coated onto a microplate. Standards or samples are premixed with biotin-labeled antibody and then pipetted into the wells. Human Anti-Monkeypox virus IgG (A29L) in the sample competitively binds to the pre-coated protein with biotin-labeled Human Anti-Monkeypox virus IgG (A29L). After washing away any unbound substances, Streptavidin-HRP is added to the wells. Following a wash to remove any unbound enzyme reagent, a substrate solution is added to the wells and color develops in inversely proportion to the amount of Human Anti-Monkeypox virus IgG (A29L) bound in the initial step. The color development is stopped and the intensity of the color is measured.

PRECISION

Intra-Assay Precision (Precision within an assay): <20% Three samples of known concentration were tested sixteen times on one plate to assess intra-assay precision. Inter-Assay Precision (Precision between assays): <20% Three samples of known concentration were tested in twenty four separate assays to assess inter-assay precision.

Sample		1	2	3	1	2	3
n		16	16	16	24	24	24
Mean (ng/mL)		16282.8	4176.8	1072.5	16037.3	4253.0	1036.4
Standard deviation		1668.9	471.1	143.4	1661.9	466.5	213.8
CV \(%\)		10.2	11.3	13.4	10.4	11.0	20.6

RECOVERY

The recovery of Human Anti-Monkeypox virus IgG (A29L) spiked to three different levels in human serum samples throughout the range of the assay was evaluated.

Sample	Dilution factor	Average (%)	Range (%)
Human serum (n=4)	1:10	97	76-121
	1:20	96	78-115

LINEARITY

To assess the linearity of the assay, human serum samples were spiked with high concentrations of Human Anti-Monkeypox virus IgG (A29L) and diluted with the Standard Diluent to produce samples with values within the dynamic range of the assay.

Sample	Dilution factor	Average (%)	Range (%)
Human serum (n=4)	1:2	100	94-112
	1:4	92	86-96
	1:8	86	85-87
	1:16	92	89-94