Birdwhistell Kate, Basinger Robert, Hayes Brian, Norton Natalie, Hurley David J, Franklin Samuel P
Department of Small Animal Medicine and Surgery (Birdwhistell, Basinger, Hayes, Franklin), University of Georgia, College of Veterinary Medicine, Athens, GA.
Regenerative Bioscience Center (Franklin), University of Georgia, College of Veterinary Medicine, Athens, GA.
J Vet Diagn Invest. 2017 Mar;29(2):143-147. doi: 10.1177/1040638717690186. Epub 2017 Feb 8.
Platelet-rich plasma has been studied extensively in dogs, but validation of enzyme-linked immunosorbent assays (ELISAs) for quantifying anabolic growth factors and inflammatory cytokines in canine plasma prepared with citrate-based anticoagulants is not available. We performed a validation of commercial ELISAs for transforming growth factor-beta 1 (TGF-β1), platelet-derived growth factor-BB (PDGF-BB), vascular endothelial growth factor (VEGF), tumor necrosis factor-alpha (TNF-α), and interleukin-1 beta (IL-1β) for use with canine plasma prepared with acid-citrate-dextrose, solution A (ACD-A). Platelet-poor plasma (PPP) anticoagulated with ACD-A as well as PPP anticoagulated with ACD-A and spiked with the relevant canine recombinant proteins were evaluated with each ELISA to calculate the efficiency of spike recovery. Replicates of the spiked PPP were also assessed in 2 additional assays to quantify intra-assay and interassay precision. The efficiency of spike recovery was within 75-125% of the expected concentration for the TGF-β1, PDGF-BB, and VEGF ELISAs. The intra- and interassay variability were <25% for the TGF-β1, PDGF-BB, VEGF, and TNF-α ELISAs. The TGF-β1, PDGF-BB, and VEGF ELISAs demonstrate acceptable efficiency of spike recovery and intra- and interassay variability, whereas the TNF-α and IL-1β ELISAs did not meet industry standards of performance with ACD-A anticoagulated canine plasma.
富含血小板血浆已在犬类中得到广泛研究，但对于用柠檬酸盐抗凝剂制备的犬血浆中用于定量合成代谢生长因子和炎性细胞因子的酶联免疫吸附测定（ELISA），尚无验证。我们对用于转化生长因子-β1（TGF-β1）、血小板衍生生长因子-BB（PDGF-BB）、血管内皮生长因子（VEGF）、肿瘤坏死因子-α（TNF-α）和白细胞介素-1β（IL-1β）的商业ELISA进行了验证，以用于用酸-柠檬酸-葡萄糖溶液A（ACD-A）制备的犬血浆。用ACD-A抗凝的乏血小板血浆（PPP）以及用ACD-A抗凝并添加相关犬重组蛋白的PPP，通过每种ELISA进行评估以计算加标回收率。加标的PPP重复样品还在另外两种测定中进行评估，以量化批内和批间精密度。TGF-β1、PDGF-BB和VEGF ELISA的加标回收率在预期浓度的75-125%范围内。TGF-β1、PDGF-BB、VEGF和TNF-α ELISA的批内和批间变异均<25%。TGF-β1、PDGF-BB和VEGF ELISA显示出可接受的加标回收率以及批内和批间变异，而TNF-α和IL-1β ELISA在使用ACD-A抗凝的犬血浆时未达到行业性能标准。
