Quantification of BSA in Tissue Engineered Medical Products by ELISA — Problems and Solutions
Aim In this study, the adaptability of an ELISA kit for quantification the residual BSA in TEMPs, the influence of rinsing protocol on reducing the residual BSA in TEMPs and the effectiveness of ultra-filtration on reducing the matrix effects of TEMPs immersion on BSA quantitative by ELISA were discussed. Methods Three kinds of TEMPs used in this study were: tissue engineered skin (TES), recombination human acellular dermal matrix (rhADM) and combination chitosan tissue engineered skin (cC-TES). The devices were rinsed according to the Directions for Use firstly. To investigating the influence of rinsing protocol on reducing the residual BSA in TEMPs, TES were rinsed by two different protocols separately. Then TEMPs immersions were prepared according to ISO10993.12, physiological saline (NS) was used as immersion medium. BSA concentration in immersions and filtrate were determined by using the “Quantitative measure of residual BSA ELISA kit” (detection range was 12.5-200ng/mL, manufactured by WUXI BOSHENG MEDICAL BIO-TEC DEVELOPMENT CO., LTD. As suspected to have some matrix effect on BSA quantification by ELISA kit, rhADM immersion was ultra-filtrated before detection. Results The results showed good correlation between dilution factors and the A450nm of TES and cC-TES immersions, correlation coefficient (r) was 0.9943±0.0007 and -0.9835±0.0037, respectively. No significant effect on BSA detection was found when NS was used as immersion medium. Comparing the results of protocol 1 and 2, the A450nm of TES immersion was significantly decreased after rising by protocol 2. After ultra-filtration, the correlations between absorption and dilution factors of rhADM immersion were improved significantly; the correlation coefficient (r) was raised from -0.7264±0.0089 to -0.9606±0.0039. Conclusions The quantitative ELISA kit was considered to be adaptability for detect the BSA in TEMPs. Different rinsing protocol may obviously affect on reducing the residual BSA in TEMPs. The matrix effects of rhADM immersion can be reduced obviously by using ultrafiltration.
Zhong Wei Gu, Yafang Han, Fu Sheng Pan, Xitao Wang, Duan Weng and Shaoxiong Zhou
Y. Fang et al., "Quantification of BSA in Tissue Engineered Medical Products by ELISA — Problems and Solutions ", Materials Science Forum, Vols. 610-613, pp. 1076-1081, 2009