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Custom Antibody Service FAQ
Western Immunoblotting Protocol (Primary Ab Incubation In BSA)
Immunohistochemistry Protocol (Paraffin)
Immunofluorescence Protocol
Flow Cytometry Protocol for Intracellular Staining Using Conjugated Secondary Antibodies
Immunohistochemistry Troubleshooting
Troubleshooting Immunofluorescence Assays
Western Blotting Troubleshooting
[more...]
For Western blots, incubate membrane with diluted antibody in 5% w/v BSA, 1X TBS, 0.1% Tween-20 at 4°C with gentle shaking, overnight.
A. Solutions and Reagents
NOTE: Prepare solutions with Milli-Q or equivalently purified water.
1. 1X Phosphate Buffered Saline (PBS) [more...]
IMPORTANT: See product data sheet for the appropriate wash buffer and antigen unmasking procedure.
• For Citrate/PBST, use steps 5a, 6a and C1.
• For Citrate/TBST, use steps 5b, 6a and C1.
• For EDTA/PBST, use steps 5a, 6b and C2. [more...]
IMPORTANT: Please refer to the APPLICATIONS section on the data sheet to determine IF THIS PRODUCT is validated and approved for the specific protocol you will be using.
A. Solutions and Reagents
NOTE: Prepare solutions with Milli-Q or equivalently purified water. [more...]
A. Solutions and Reagents
1. 1X Phosphate Buffered Saline (PBS): Dissolve 8 g NaCl, 0.2 g KCl, 1.44 g Na2HPO4 and 0.24 g KH2PO4 in 800 mL distilled water (dH2O). Adjust the pH to 7.4 with HCl and the volume to 1 liter. Store at room temperature.
2. Formaldehyde (methanol free) [more...]
Weak or No Staining
Sources
Inadequate deparaffinization
Inactive primary antibodies [more...]
Problem: Positive Control is negative or very weak
Possible Causes:
1. Microscope must be equipped with a filter system for FITC (maximum excitation wavelength 490 nm, mean emission wavelength 530 nm) and 400X magnification. [more...]
Problem: Poor transfer quality: The poor transfer of proteins to the membrane can be visualized by staining the gel after transfer. [more...]
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- Westem Immunblotting Protocol(Primary Ab Incubation InBSA)
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