Human TIM-1/KIM-1/HAVCR DuoSet ELISA

Protocol tips

Upstream tips	Protocol tips	Downstream tips
-Bring all reagents to room temperature before use. Allow all components to sit for a minimum of 15 minutes with gentle agitation after initial reconstitution. -Working dilutions should be prepared and used immediately.	-A standard curve should be generated for each set of samples assayed. -It is suggested to start Reagent Diluent optimization for serum and plasma samples by using PBS supplemented with 10-50% animal serum. -Do not use buffers with animal serum to reconstitute or dilute the Detection Antibody or Streptavidin-HRP B.

Upstream tips
-Bring all reagents to room temperature before use. Allow all components to sit for a minimum of 15 minutes with gentle agitation after initial reconstitution. -Working dilutions should be prepared and used immediately.
Protocol tips
-A standard curve should be generated for each set of samples assayed. -It is suggested to start Reagent Diluent optimization for serum and plasma samples by using PBS supplemented with 10-50% animal serum. -Do not use buffers with animal serum to reconstitute or dilute the Detection Antibody or Streptavidin-HRP B.

Publication protocol

Neutrophil gelatinase-associated lipocalin (NGAL), tissue inhibitor metalloproteinase type 2 and insulin-like growth factor-binding protein type 7 (TIMP-2*IGFBP-7), kidney injury molecule 1 (KIM-1), interleukin-18 (IL-18), and liver-type fatty acid-binding protein (L-FABP) were measured on the urine samples collected at baseline (prior to CPB initiation) and then at 2, 6, 12, 24, 48, and 72 hours following CPB initiation. TIMP-2*IGFBP-7 was measured using the Astute® 140 meter (Astute Medical, Inc., San Diego, California). The authors have previously reported the cut-off points of TIMP-2*IGFBP-7 associated with AKI [20]. Urine NGAL was measured using a commercially available assay (NGAL ELISA Kit 036; Bioporto, Grusbakken, Denmark) that specifically detects human NGAL [26]. The intra-assay CV was 2.1% and inter-assay variation was 9.1%. Urine IL-18 (Kit 7625) and L-FABP (Kit Z-001) were measured using commercially available ELISA kits (Medical & Biological Laboratories Co., Nagoya, Japan and CMIC Co., Tokyo, Japan, respectively) per manufacturer’s instructions. Intra-assay CV for IL-18 and L-FABP were 7.3% and 6.1% respectively. Interassay CV were 7.5% and 10.9% for IL-18 and L-FABP respectively. The urine KIM-1 ELISA was performed using validated commercially available reagents (Duoset DY1750, R & D Systems, Inc., Minneapolis, MN) as previously described.

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Manufacturer protocol

Download the product protocol from R&D Systems for Human TIM-1/KIM-1/HAVCR DuoSet ELISA below.

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