((Running title – jobm.201400330))
Simple viral plaque assay for Enterovirus
Method Paper
A simple and highly repeatable viral plaque assay for Enterovirus 71
Yingxian Yin, Yi Xu, Zhiying Ou, Ling Su and Huimin Xia
Institute of Pediatrics, Guangzhou Women and Children’s Medical Center, Guangzhou,
China
DOI 10.1002/jobm.201400330
Table S1. Comparison of new plaque assay with other existing procedures.
New
overlay
plate
fixation
staining
PA
PB
PC
PD
[1]
[2]
[3]
[4]
Avicela,b
+
+
-
-
-
Agarose
Methylcellulose
6-well
24-well
96-wellb
Formaldehyde in
Saline
Formaldehyde in
MEM
Direct
fixation
without removing
overlayc
Immunostainingd
Neural red
Crystal violet
+
+
+
+
+
+
+
-
+
+
+
+
+
+
-
+
+
+
+
-
+
-
-
+
+
-
+
-
-
+
-
+
+
+
-
+
-
+
+
Note: New, the new procedure in this report. PA, PB, PC and PD， existing procedures in literature.
asimple
preparation; bpotential high throughput and more economic; chigh repeatability; dexpensive; + step
or material involved; - step or material not involved.
References
[1] Matrosovich, M., Matrosovich, T., Garten, W., Klenk, H.D., 2006. New low-viscosity
overlay medium for viral plaque assays. Virol. J., 3, 63.
[2] McKimm-Breschkin, J.L., 2004. A simplified plaque assay for respiratory syncytial
virus--direct visualization of plaques without immunostaining. J. Virol. Methods, 120(1),
113-117.
[3] Dulbecco, R., 1952. Production of plaques in monolayer tissue cultures by single particles
of an animal virus. Proc. Natl. Acad. Sci. USA, 38 (8), 747-752.
[4] Wang, S.M., Chen, I.C., Su, L.Y., Huang, K.J. et al., 2010. Enterovirus 71 infection of
monocytes with antibody-dependent enhancement. Clin. Vaccine Immunol., 17(10),
1517-1523.
Figure S1. Natural sedimentation after serial dilution of Avicel (R-591) overlay media.
Labeling: R-591 was dispersed at 37 ℃ for 1 h. From left to right, the concentration was 2.4%
(A, in distilled water), 1.2% (B), 0.6% (C), 0.3% (D) and 0.15% (E), respectively. Except for
the initial concentration (A), all others were dispersed in 1×DMEM (red fluid).
Figure S2. HSV-1 plaque assay on Vero cell monolayer in 24-well plate by new procedure.