TROUBLESHOOTING
Troubleshooting advice can be found in Table 1.
Troubleshooting table
Problem
Possible reason
Solution
Step 8: the low sensitivity of
The operation time of step 7 is
Centrifuge immediately.
slides by scanner
too long
Step 8: the low fluorescence
The pH of blocking/ incubation
image of lectin microarray
buffer is not proper
Adjust pH value to 7.4
The low amount of Cy3-labeled
Add the amount of Cy3-labeled
protein
protein
The temperature of blocking and
Adjust the temperature to 25 ºC
incubation is not proper
The incubation time is not
Adjust the incubation time to
proper
2-3h
Step 37: a failure of silver
The time of step 31 is not
At least 2 h
staining
enough
The serial of fluid is not fresh
Sensitization
fluid,
solution
developer
and
staining
had
better prepare when you use
Step 44: Step 44: a failure of
lectin blotting
Washing is not enough
To add an extra several times
The gel is polluted by other
Avoid
proteins
hands , contaminated utensil)
The transfer buffer is not fresh
The transfer buffer had better
other
pollution
prepare when you use
(e.g.
Washing is not enough
The
PVDF
membrane
To add an extra several times
is
Avoid
other
pollution
(e.g.
polluted by other proteins
hands, contaminated utensil)
The blocking time is not proper
Adjust the blocking time up to
2h at room temperature
The low amount of Cy5-labeled
Add the amount of Cy5-labeled
protein
protein
The temperature of blocking and
Adjust the temperature to 4 ºC
incubation is not proper
in the dark
The incubation time is not
Adjust the incubation time up to
proper
10h
Figure legends
Figure 1. Relative expression levels of the terminal Siaα2-6Gal recognized by SNA and Siaα2-3Gal recognized by
MAL-II in human saliva using lectin microarrays. MH: male healthy groups, FH: female healthy groups, MD:
male T2DM patients, FD: female T2DM patients
Figure 2. Validation of the terminal Siaα2-6Gal recognized by SNA and Siaα2-3Gal recognized by MAL-II in all
salivary groups. The summed fluorescence intensities (SFIs) of the bands marked by red lines at approximately 25
kDa for all salivary groups from lane 1 to lane 4 were read by ImageQuant Tools. (A) Lane 1: male healthy group
(MH); lane 2: female healthy group (FH); lane 3: male T2DM patients (MD); lane 4: female T2DM patients (FD).
(B) The summed fluorescence intensities for SNA and MAL-II strains binding to samples from the T2DM salivary
group.
Figue 1
Figure 2