Human c peptide elisa

For the human islet xenograft model, 1,000 human IEQ from cadaveric donors were transplanted under the renal capsule of anesthetized atleast 8 weeks old male NOD/LtSz-scid IL2R gamma null NSG Tg(RIP-HuDTR) mice. Human C-peptide levels were monitored on day 7, day 14, and day 21 post-transplantation using a Human C-peptide ELISA (ALPCO Diagnostics, Salem, NH) to ensure successful islet engraftment.
The animal care, handling, and all animal studies were performed following ARRIVE guidelines, in compliance with, and with approval of, the Oregon Health & Science University's Institutional Animal Care and Use Committee (IACUC, protocol# IP00001290).

Samples (aggregated FACS-sorted β cells or hESC-derived spheres) were removed from media and equilibrated by incubation for 30 min at 37 °C in KRB+2.8 mM glucose with or without 30 mM ivabradine (Sigma-Aldrich). Cells were then transferred to fresh assay media (KRB+2.8 mM (basal) or 22.2 mM (stimulated) glucose with or without ivabradine for 1.5 h at 37 °C). After incubation, buffers were collected and frozen for subsequent human c-peptide-specific ELISA analysis (Mercodia). For hESC-derived spheres, total human c-peptide content analysis was performed by measurement of an aliquot of acidic ethanol lysed clusters by human c-peptide ELISA (Alpco). Statistical analyses of the results were performed using Graphpad Prism 4.0 (for analysis of variance tests) and Microsoft Excel (for mean, standard deviation and t-tests).