This investigation analyzed, in a cross-sectional study, the possible relationship between gingival crevicular fluid (GCF) elastase-like protease (ELP) levels and the periodontal clinical parameters or the presence of specific bacteria in subgingival plaque. A total of 388 periodontal sites from 8 adult periodontitis patients were examined for plaque index (PII), gingival index (GI), pocket depth (PD) and alveolar bone loss (ABL). GCF ELP levels were determined as ELP alpha-1 protease inhibitor (ELP-alpha 1-PI) complex levels with a commercially available ELISA. Subgingival plaque samples were tested for the presence of Bacteroides gingivalis, B. intermedius and Actinobacillus actinomycetemcomitans by indirect immunofluorescence (IF) microscopy. GCF ELP-alpha 1-PI levels were then correlated with clinical periodontal indices and proportions of IF-positive bacteria per site. Statistically significant positive correlations were found between GCF ELP-alpha 1-PI concentrations and subgingival Bacteroides proportions. When the sites examined were analyzed depending on the level of each clinical parameter, the levels of these correlations changed. A. actinomycetemcomitans correlated highly (r = 0.716) with ABL for sites with low GI score. The correlations between GCF ELP-alpha 1-PI and B. gingivalis (r = 0.642) or B. intermedius (r = 0.774) were the highest for ABL less than or equal to 20% and PD less than or equal to 3 mm, respectively. The strong association between GCF ELP-alpha 1-PI concentrations and subgingival bacteria previously associated with advancing periodontitis indicates that measurement of GCF ELP-alpha 1-PI concentrations may be useful in the evaluation of periodontal sites, especially those with very little or no tissue destruction.