The 3D7 strain of P. falciparum was preserved at the Laboratory of Pathogen Infection and Immunity (Jiangnan University, Wuxi, China). The parasites were grown at 37 °C in a mixed environment of 90% N2, 5% O2, and 5% CO2, and were maintained in RPMI Medium 1640 (Gibco, New York, USA) containing O+ human erythrocytes (4% hematocrit), HEPES (Meilunbio, Dalian, China), NaHCO3 (Meilunbio, Dalian, China), AlbuMax Ⅱ (Sigma, San Francisco, CA, USA), hypoxanthine (Sigma, San Francisco, CA, USA), and gentamicin (Solarbio, Beijing, China).
The schizonts were purified by 60% percoll (Solarbio, Beijing, China) gradient centrifugation, and lysed in 0.1% saponin (diluted in PBS) for 5 min on ice with intermittent mixing. The lysed material was centrifuged at 15,000× g for 5 min and washed thrice with PBS. Then, the parasite lysate was collected and boiled in a SDS-PAGE sample loading buffer (Meilunbio, Dalian, China) for 7 min [13 (link)]. The total protein was separated on SDS-PAGE gel, and native PfSRA in the P. falciparum crude protein was captured by the anti-rPfSRA mice sera at 1:1000 dilution overnight and detected by HRP-conjugated goat anti-mouse IgG (CWBio, Beijing, China).
The schizonts were purified by 60% percoll (Solarbio, Beijing, China) gradient centrifugation, and lysed in 0.1% saponin (diluted in PBS) for 5 min on ice with intermittent mixing. The lysed material was centrifuged at 15,000× g for 5 min and washed thrice with PBS. Then, the parasite lysate was collected and boiled in a SDS-PAGE sample loading buffer (Meilunbio, Dalian, China) for 7 min [13 (link)]. The total protein was separated on SDS-PAGE gel, and native PfSRA in the P. falciparum crude protein was captured by the anti-rPfSRA mice sera at 1:1000 dilution overnight and detected by HRP-conjugated goat anti-mouse IgG (CWBio, Beijing, China).
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