ELISpot Homology and Cross-Reactivity Results
Due to the high degree of similarity observed between human and non-human primate cytokine protein sequences (See Table 1), R&D Systems has evaluated the potential use of our human ELISpot kits to detect cytokines from non-human primate cells. Cells from three different species of monkey were tested with each of our human ELISpot kits and found to cross-react with select analytes (See Table 2). Not all species represented in the table have been tested at this time.
| Human Protein Accession Number | ||||||
| Table 1. Amino acid sequences (note: protein accession number) for several human cytokines were BLASTed for homology to non-human primate protein sequences. | ||||||
| Species | IL-1 beta NP_000567 | IL-2 NP_000577 | IL-4 NP_000580 | IL-6 NP_000591 | IL-13 NP_002179 | TNF-alpha NP_000585 |
| Rhesus monkey (Macaca mulatta) | 91% | 94% | 93% | 96% | NA | 95% |
| Pigtail monkey (Macaca nemestrina) | 91% | 94% | NA | NA | NA | NA |
| Cynomolgus monkey (Macaca fasicularis) | 90% | 93% | 92% | 96% | NA | 94% |
| Common squirrel monkey (Saimiri sciureus) | 89% | 89% | NA | 87% | NA | 86% |
| Mangabey (Cercocebus torquatus) | 91% | 94% | 92% | 95% | NA | NA |
| Baboon (Papio hamadryas) | NA | 90% | NA | NA | NA | 95% |
| ELISpot Kit1 | ||||
| Table 2. Peripheral blood mononuclear cells from each of the listed species were stimulated with 50 ng/mL of phorbol 12-myristate-13-acetate and 0.5 µg/mL of calcium ionomycin (IL-2, IL-13) or 3 µg/mL phytohemagglutinin (PHA) (IL-4, IL-6) overnight at 37° C in a 5% CO2 incubator. The data shown is representative data from four independent experiments. Due to the inherent variability observed in immune responses within a group of subjects, assay results may vary. | ||||
| Species | Human IL-2 (5x105 cells/mL) | Human IL-4 (1x106 cells/mL) | Human IL-6 (1x105 cells/mL) | Human IL-13 (5x106 cells/mL) |
| Spots/well | Spots/well | Spots/well | Spots/well | |
| Rhesus monkey (Macaca mulatta) | 400 | NCR2 | 195 | 31 |
| 417 | NCR | 165 | 15 | |
| 412 | NCR | 209 | 29 | |
| 425 | NCR | 176 | 15 | |
| Pigtail monkey (Macaca nemestrina) | 412 | 36 | 52 | NCR |
| 402 | 29 | 36 | NCR | |
| 430 | 28 | 30 | NCR | |
| 462 | 42 | 27 | NCR | |
| Cynomolgus monkey (Macaca fasicularis) | Confluent3 | 36 | 42 | 73 |
| Confluent | 28 | 37 | 61 | |
| Confluent | 40 | 30 | 62 | |
| Confluent | 25 | 46 | 63 | |
Notes:
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