DESCRIPTION

Used for the quantitative determination of Anti-Infliximab concentration in serum and plasma.

PRINCIPLE OF THE ASSAY

This assay employs the quantitative sandwich enzyme immunoassay technique. Infliximab has been pre-coated onto a microplate. Samples or standards are pipetted into microwells and Anti-Infliximab will be captured by immobilized Infliximab. After washing away any unbound substances, a biotin-labeled Infliximab is added to the wells. 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 proportion to the amount of Anti-Infliximab 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): <10% Three samples of known concentration were tested sixteen times on one plate to assess intra-assay precision. Inter-Assay Precision (Precision between assays): <15% 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)		241.6	58.4	9.4	235.6	48.7	9.9
Standard deviation		8.1	1.8	0.9	14.2	1.8	0.6
CV \(%\)		3.4	3.0	9.9	6.0	3.7	6.4

RECOVERY

The recovery of Anti-Infliximab 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=3)	1:10	92	88-97
	1:20	92	86-97

LINEARITY

To assess the linearity of the assay, human serum samples were spiked with high concentrations of Anti-Infliximab 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	101	98-102
	1:4	103	101-105
	1:8	104	100-107
	1:16	94	80-101