Depositors Notes: Suitable to use in immunostaining and western blots. 1/ Staining of embryos and discs; both fixative 1 (3.7% formaldehyde in PBS) and fixative 2 (4% paraformaldehyde in CMF & GBS containing 1mM CaCl2 (see details in Dev. Biol. 165, 716-726)) work. 2/ Staining of S2 cells; fixative 1 doesn't work, use fixative 2. 3/ Staining of pupal or adult heads for light microscope; find details in our MCN paper and references therein such as Uchida et al. (4% paraformaldehyde-based fixative on ice for 1-2 hr) and Hoshino et al. (PLP-based fixative on ice for 1-2 hr). For better fixation, we cut pupal or adult heads into halves. 4? ImmunoEM of adult heads; find details in our MCN paper. We do not add EDTA or EGTA in any fixatives, because cadherins are easily degraded under Ca++- free conditions.
Recommended Applications: Immunofluorescence, Immunohistochemistry, Western Blot
These hybridomas were created by your colleagues. Please acknowledge the hybridoma contributor and the Developmental Studies Hybridoma Bank (DSHB) in the Materials and Methods of your publications. Please email the citation to us.
For your Materials & Methods section:
DN-Ex #8 was deposited to the DSHB by Uemura, T. (DSHB Hybridoma Product DN-Ex #8)
Storage and Handling Recommendations
Although many cell products are maintained at 4°C for years without loss of activity, shelf-life at 4°C is highly variable.
To ensure retention of antibody activity, we recommend aliquotting the product into two parts:
1) a volume of antibody stored at 4°C to be used within two weeks.
2) the remaining product diluted with an equal volume of molecular grade glycerol and stored at -20°C.
While optimal Ig concentration for an application will vary, a good starting concentration for immunohistochemistry (IHC),
immunofluorescence(IF) and staining is 2-5 µg/ml.
For Western blots, the concentration is decreased by one order of magnitude (that is, 0.2-0.5 µg/ml).
All cell products contain the antimicrobial ProClin. See our FAQ section for additional information.