The objective of this study was to evaluate the relationship between integrated connective tissue (ICT), that is, the presence of connective tissue into the membrane structure, and the clinical outcome of membrane-supported periodontal surgery. Twenty-six systemically healthy subjects affected by chronic adult periodontitis were enrolled in the study. One tooth site per patient, associated with an angular bony defect and an attachment loss of > 7 mm, was selected to be treated by means of a guided tissue regeneration procedure using a bioabsorbable membrane. Barrier material was surgically removed after 4 weeks for SEM analysis. For each treated site, the difference in clinical attachment loss, probing depth, and gingival recession between the baseline examination and follow-up 6 months after the second surgery was calculated. Gain of attachment was statistically (P < 0.001) greater in sites with no membrane exposure when compared to sites with exposed barrier material (5.5 +/- 1.0 vs. 4.0 +/- 0.6), while further gingival recession was greater (3.0 +/- 0.9 vs. 2.1 +/- 0.5) in sites with clinically exposed membranes. The results of SEM analysis revealed that when connective tissue structures were observed on membrane surfaces, no bacteria could be detected; conversely, areas heavily colonized by bacteria did not show the presence of connective tissue. Regression analysis indicated that integrated connective tissue on the external layer of the barrier material was positively correlated with the amount of attachment gain and negatively with the amount of gingival recession. Bacterial colonization of the membrane was negatively correlated with attachment gain and positively with gingival recession. It was concluded that connective tissue integration is an important biological phenomenon in preventing membrane exposure and bacterial plaque colonization and thus in enhancing the clinical outcome following guided tissue regeneration surgery.