Isolation and purification of pulmonary surfactant-associated protein A from human amniotic fluid and its bioactivity assessment.
OBJECTIVE: To isolate and purify pulmonary surfactant-associated protein A(SP-A) from human amniotic fluid. METHODS: Human amniotic fluid was collected from 11 parturients and SP-A was isolated by extraction and dialysis followed by purification procedures. The specificity of SP-A was detected by Western blotting and the bioactivity assessed by MTP-1 film balance. RESULTS: An amount of 12 mg (20 ml) purified SP-A was obtained, and the protein presented a single band upon SDS-PAGE with relative molecular mass of 31 000. Western blotting also displayed one specific band. Film balance demonstrated that the minimum surface tension alphamin of Exosurf, an synthetic pulmonary surfactant, was 7.7+/-0.3, significantly higher than alphamin of Exosurf and SP-A mixture(1.5+/-0.22, P<0.05), indicating enhanced bioactivity of the mixture. CONCLUSION: Dialysis can effectively purify SP-A from human amniotic fluid with little damage of its biological activity.
Liu
Yong-Mei
YM
Department of Pediatrics, Zhujiang Hospital, First Military Medical University, Guangzhou 510282, China.
Feng
Zhi-Chun
ZC
Wu
Bing-Yi
BY
Du
Jiang
J