DESCRIPTION

Aminoacyl-tRNA synthetases catalyze the aminoacylation of tRNA by their cognate amino acid. Because of their central role in linking amino acids with nucleotide triplets contained in tRNAS, aminoacyl-tRNA synthetases are thought to be among the first proteins that appeared in evolution. Two forms of isoleucine-tRNA synthetase exist, a cytoplasmic form and a mitochondrial form. This gene encodes the mitochondrial isoleucine-tRNA synthetase which belongs to the class-I aminoacyl-tRNA synthetase family.

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	6.25	0.14	2.29
2	8	1.62	0.07	4.04
3	8	0.91	0.05	5.58

Inter-assay Precision

Sample	n	Mean (ng/mL)	SD	CV%
1	16	6.13	0.38	6.26
2	16	1.69	0.10	5.84
3	16	0.95	0.05	5.37

RECOVERY

The recovery of human/rat IARS2 spiked to three different levels throughout the range of the assay in lysate was evaluated.

Sample Type		Average% of Expected	Range (%)
Lysate	1:8	81	80-81
	1:16	85	78-102

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 lysate was initially diluted 1:2. )

		Lysate
1:2	Average% of Expected	100
	Range (%)	-
1:4	Average% of Expected	87
	Range (%)	86-88
1:8	Average% of Expected	82
	Range (%)	82-83
1:16	Average% of Expected	84
	Range (%)	77-91

CALIBRATION

This immunoassay is calibrated against a highly purified recombinant human/rat IARS2 produced at Sunlong Medical™.