Fig 1.
Optimization of KGH P protein expression in BL21(DE3) E. coli and purification.
(A) Protein expression according to temperature. M, protein size marker; 1, uninduced total cell lysate; 2 and 6, total cell lysate prepared from cells induced with 0.4 mM IPTG using 20 mM Tris lysis buffer; 3 and 7, total cell lysate prepared from cells induced with 0.4 mM IPTG using 50 mM NaH2PO4lysis buffer; 4 and 8, cell lysate supernatant prepared from cells induced with 0.4 mM IPTG using 20 mM Tris lysis buffer; 5 and 9, cell lysate supernatant prepared from cells induced with 0.4 mM IPTG using 50 mM NaH2PO4lysis buffer. (B) Protein expression according to buffer composition. M, protein size marker; 1 and 8, cell lysate supernatant; 2 and 9, flow-through; 3, 4, 10, 11, wash; 5–7, 12–14, elution.
Fig 2.
IFA reactivity according to RABV strain and infected cell type.
(A) BHK-21 cells and (B) N2a cells. The cells were infected with the KGH, CVS-11, and ERA strains for 48 h. The results were observed under a fluorescence microscope at a magnification 400×.
Fig 3.
Comparison of the detection limits of the 16B8-Alexa diagnostic antibody with that of the DFA.
(A) 16B8-Alexa and (B) commercial DFA reagent were five-fold serially diluted in PBS containing 0.00025% Evans blue to dilutions ranging from 1:100 to 1:12,500. Fluorescence microscopic images are shown at a magnification of 400×.
Table 1.
RFFIT results using DFA reagent and 16B8-Alexa.
Fig 4.
Correlation of RFFIT results of 414 clinical specimens, using DFA reagent and 16B8-Alexa.
Significant correlation was observed (r = 0.995, p<0.001).