Protein Protocols Protein Protocols

376 Alba and Daban
Fig. 1. According to Weigele et al. (2), the reaction of MDPF (A) with primary amino groups
of proteins produces the fluorescent adduct (B); the excess reagent is hydrolyzed forming the
nonfluorescent product (C).
Fig. 2. (A) Example of MDPF staining of different proteins on PVDF membranes. The blot
was equilibrated twice (5 min each time) in borate buffer and stained for 10 min with MDPF
(see Subheading 3.). Proteins, from top to bottom, are: BSA, ovalbumin, glyceraldehyde-3phosphate
dehydrogenase, β-lactoglobulin, and α-lactalbumin; the amount of each protein
loaded initially onto the SDS gel before electroblotting was 100 (left lane) and 50 (right
lane) ng. (B) After staining with MDPF ovalbumin was immunodetected with the ECL system
(Amersham).
3. Transfer apparatus (e.g., Mini-Trans-Blot Cell [Bio-Rad]).
4. Opaque plastic box for membrane equilibration and staining.
5. Orbital shaker.
6. Transilluminator equipped with midrange ultraviolet (UV) bulbs (~300 nm) to excite
MDPF labeled proteins on blots (e.g., Foto UV 300 [Fotodyne Inc., Harland, WI] or the