Gdv programmable mpt reader dv 990bv4

Serum ET-1 was assessed using a specie-specific commercial sandwich ELISA kit (Claud-Clone Corp, Katy, TX, USA). The detection range was 6.17–500 pg/ml, and intra- and interassay CV were <10 and <12%, respectively. Absorbance was determined using a microplate reader (GDV programmable MPT Reader DV 990BV4, Agilent Technologies, Santa Clara, CA, USA) at 450 nm.
The serum troponin I levels were assessed using a human commercial kit [ADVIA Centaur High Sensitivity Troponin I (TNIH)], previously validated in canine species (52 (link)). The detection limit was 0.006 ng/ml. The analyses were performed using a chemiluminescence immunodiagnosis system (IMMULITE 1000 Systems, Siemens Healthcare S.r.l., Milan, Italy).

Serum insulin analysis was performed for each sample using an ELISA kit (canine insulin ELISA, Mercodia AB). The insulin/glucose ratio (I/G) was calculated as the serum insulin (μU/ml) × 100/serum glucose (mg/dl), according to Bailhache et al. (51 (link)). In particular, the detection range was 2.3–173 mU/L with a detection limit of 1.15 mU/L. Absorbance was determined using a microplate reader (GDV programmable MPT Reader DV 990BV4, Agilent Technologies, Santa Clara, CA, USA) at 450 nm.
Serum leptin was estimated by an ELISA kit (Canine Leptin ELISA Cat. EZCL-31K, Millipore, Billerica, MA, USA). The detection limit was 0.2 ng/L, and intra- and interassay coefficients of variation (CV) were <5%. Absorbance was determined by a spectrophotometer with a wavelength of 450 nm (Epoch, BioTek Instruments Inc., Winooski, VT, USA).

Serum TNF-α and IL-6 were measured by canine cytokine ELISA kit (Genorise, Glen Mills, PA, USA). TNF-α detection range assay was 1–2,200 pg/ml with intra- and interassay CV <7 and <9%, respectively. IL-6 detection range assay was 50–3,200 pg/ml with intra- and interassay CV <6 and <9%, respectively. Absorbance was determined using a microplate reader (GDV programmable MPT Reader DV 990BV4, Agilent Technologies, Santa Clara, CA, USA) at 450 nm.