Expression of H3N2 nucleoprotein in maize seeds and immunogenicity in mouse
Hartinio N Nahampun1,2, Brad Bosworth3, Joan Cunnick3, Mark Mogler4, Kan Wang2,*
1
Interdepartmental Plant Biology Major, Iowa State University, Ames, IA 50011-1010 USA
2
Department of Agronomy, Iowa State University, Ames, IA 50011-1010, USA
3
Department of Animal Science, Iowa State University, Ames, IA 50011-1010 USA
4
Harrisvaccines, Inc, Ames, IA 50010 USA
*To whom correspondence should be addressed: E-mail: kanwang@iastate.edu
Supplemental Materials
Maize-derived NP (MNP) oral feeding experiment
Female BALB/c mice (6-8 weeks old) were purchased from Charles River Laboratories
International Inc. (Willmington, MA). The experiments were approved by Institutional Animal
Care and Use Committee (IACUC) of Iowa State University. All mice were maintained under
pathogen-free conditions and treated humanely. Mice were housed in plastic cage (3 mice per
cage) at 20°C with reverse 12 h light/dark cycle. The animals were allowed to 2 weeks
adjustment before the experiment started. Bedding was changed twice per week and mice had
ad-libitum access to feed and water. Body weight was recorded every week. Unless otherwise
stated, specimen collection, antibody analysis and ELISPOT was conducted in the same manner
of MNP oral administration study.
Immunization
The mouse oral feeding experiment on MNP was conducted as follow. Six groups consisted of 6
mice per group were fed with three doses of maize pellet (except group 5 that received VNP
subcutaneously) in prime-boost design (Day 0, 21 and 42) as summarized in Table S1. In the
groups receiving MNP and VNP, maize pellets were adjuvanted with or without MLB (maize
derived LT-B). Mice were fasted 12 hours before immunization, normal feeding was taken away
except water. During the day of immunization, each mouse was put in a single container and one
maize pellet was given. Mice receiving subcutaneous injection accepted the same treatment as
well. Mouse was let to finish the pellet for 1 hour then all mice were put back in their cage and
normal feed and water was put back.
1
Supplemental results
Table S1. MNP oral feeding experiment in mouse
Groups
# mouse
Immunogens
1
2
3
4
5
6
6
6
6
6
6
6
MNP
MNP + MLB
VNP + NTM
VNP + MLB
VNP
NTM
Prime/boost
pellet
pellet
pellet
pellet
subcutaneous
pellet
20 µg
20 µg
20 µg
20 µg
20 µg
20 µg
Fig. S1 NP specific antibody in experimental mouse serum and fecal after oral feeding
experiment with MNP
Mouse fecal IgG
1.20
Mouse sera IgG
1.50
*
0.80
0.60
S/N ratio
S/N ratio
1.00
0.40
1.00
0.50
0.20
0.00
0.00
Groups
Groups
S/N (Sample/Negative) response is the ratio of the sample optical density (OD) reading to the kit negative control OD reading. Statistical
analysis using JMP10 (SAS Inc), Turkey’s test (α < 0.05)
2