Abstract
Tumor necrosis factor alpha (TNFalpha) in human skin: a comparison of different antibodies for immunohistochemistry.
van der Laan N, de Leij LF, Buurman W, Timens W, ten Duis HJ.
Department of Surgery, University Hospital Groningen, The Netherlands.
Conflicting results have been reported regarding the localization and presence of TNFalpha in normal human skin. To study TNFalpha expression, we tested a panel of antibodies directed against human TNFalpha. First, antibodies were tested for immunoreactivity on cytospots of isolated LPS-stimulated peripheral blood mononuclear cells. Second, antibodies were tested to detect recombinant TNFalpha in Western blots. Some antibodies were found to be unable to detect recombinant TNFalpha in the blots. However, most antibodies were able to bind TNFalpha protein, but they did not bind to other irrelevant proteins that were also present in the blots. Finally, antibodies were tested on cryosections of normal human skin. Antibodies that did not react with TNFalpha in the blots were incubated with TNFalpha before the staining procedure to see whether these antibodies specifically bound TNFalpha. We found that, although all the antibodies bound TNFalpha, there were clear differences in staining patterns. This indicates that these antibodies may recognize distinct epitopes or different forms of TNFalpha. The differences found in this study and those reported previously could be the result of differences in the concentration of antibody used, the staining procedure or specificity of the antibody itself. So, for unambiguous interpretation of data, it is important to know the characteristics of the antibodies used
van der Laan N, de Leij LF, Buurman W, Timens W, ten Duis HJ.
Department of Surgery, University Hospital Groningen, The Netherlands.
Conflicting results have been reported regarding the localization and presence of TNFalpha in normal human skin. To study TNFalpha expression, we tested a panel of antibodies directed against human TNFalpha. First, antibodies were tested for immunoreactivity on cytospots of isolated LPS-stimulated peripheral blood mononuclear cells. Second, antibodies were tested to detect recombinant TNFalpha in Western blots. Some antibodies were found to be unable to detect recombinant TNFalpha in the blots. However, most antibodies were able to bind TNFalpha protein, but they did not bind to other irrelevant proteins that were also present in the blots. Finally, antibodies were tested on cryosections of normal human skin. Antibodies that did not react with TNFalpha in the blots were incubated with TNFalpha before the staining procedure to see whether these antibodies specifically bound TNFalpha. We found that, although all the antibodies bound TNFalpha, there were clear differences in staining patterns. This indicates that these antibodies may recognize distinct epitopes or different forms of TNFalpha. The differences found in this study and those reported previously could be the result of differences in the concentration of antibody used, the staining procedure or specificity of the antibody itself. So, for unambiguous interpretation of data, it is important to know the characteristics of the antibodies used
| Original language | English |
|---|---|
| Pages (from-to) | 226-232 |
| Number of pages | 6 |
| Journal | Archives of Dermatological Research |
| Volume | 293 |
| Issue number | 5 |
| DOIs | |
| Publication status | Published - 1 Jan 2001 |