Internalization and Colocalization of a Polyclonal Antibody Against Porphyromonas gingivalis FimA type I in Infected...

The aim of this work was to investigate the effect of a rabbit polyclonal antibody specific to Porphyromonas gingivalis FimA type I (FimI) protein internalized into P. gingivalis infected cells.Rabbits were immunized with P. gingivalis FimI protein and the serum was collected for immunoglobulin (Ig) purification. For visualization of the antibody inside the cells, it was labeled with Cy3 dye. Live P. gingivalis was labeled with PKH67 dye. Rabbit anti-FimI Ig-Cy3 was internalized into H357 cells infected with P. gingivalis-PKH67 by electroporation or coincubation. Location of the Ig or P. gingivalis was observed under fluorescence microscope or confocal microscope. Percentage of P. gingivalis-PKH67 infected cells was analyzed by flow cytometry.Normality of data distribution was tested by Shapiro-Wilk test. The data failed normality test and were further analyzed by Kolmogorov-Smirnov test.Rabbit anti-P. gingivalis FimI Ig-Cy3 and P. gingivalis-PKH67 were both located next to the nucleus. The rabbit anti-FimI Ig-Cy3 was able to enter H357 cells after the cells were cultured in the medium containing the labeled Ig for 16 hours. The location of the Ig was near the nucleus as found in cells electroporated with the Ig-Cy3. The percentage of P. gingivalis-PKH67 infected cells seemed to be decreased after the infected cells internalized anti-FimI Ig by electroporation. However, it was not statistically significance.Rabbit anti-P. gingivalis FimI Ig and P. gingivalis was colocalized near the nucleus. And the rabbit anti-FimI Ig was able to enter H357 cells by coincubation method.