title: Numbers and vitality of leukocytes in pocket washings of untreated periodontitis lesions in humans utilizing a novel intracrevicular lavage technique.
abstract: The purpose of the present study was to develop an intracrevicular lavage technique, and to use it for assessment of the total number and % of vital leukocytes from untreated periodontitis pockets. A lavage device was developed, consisting of specially crafted cannulas, a vacuum pump and a charge amplifier. In vivo evaluations showed that there was a linear relationship between lavage time and lavage volume; 2-6 lavage samples were obtained from each of 20 patients with untreated advanced periodontitis, and the lavage time was measured in a subsample of 16 pockets. The lavage fluid was centrifuged, and the supernatant was separated from the cellular components. The cells were vital-stained using two methods, trypanblue exclusion method (TB) and ethidium-bromide fluorescein-diacetate stain (EB-FDA). Numbers of vital and non-vital leukocytes per sample were assessed using a Neubauer chamber. The number of erythrocytes per sample was evaluated using the same counting method. The results included 95 samples obtained from the 20 patients: 76% of all samples ranged in pocket depth between 4 and 8 mm. The lavage technique provided an overall mean lavage volume of 282.37 microliters in an average time of 16.44 seconds. The mean number of leukocytes per sample was 22.20 x 10(3) (TB) and 24.48 x 10(3) (EB-FDA). Percent vital leukocytes were 72.27 (TB) and 72.63 (EB-FDA). EB-FDA had a lower counting error than TB. The low erythrocyte counts per sample suggested that subgingival bleeding during sampling was negligible. Spearman correlation coefficients showed weak associations between pocket depth and leukocyte counts, % of vital leukocytes and erythrocytes. Due to the short sampling time this new intracrevicular sampling technique permits sampling of pockets before the tissue responds to the stimulus of the lavage device, and provides subgingival washings with high numbers of leukocytes.
authors.affiliation: Abteilung für Parodontologie, Universität Bern, Schweiz.
authors.forename: U
authors.initials: U
authors.lastname: Zappa
authors.forename: G
authors.initials: G
authors.lastname: Boretti
authors.forename: H
authors.initials: H
authors.lastname: Graf
authors.forename: D
authors.initials: D
authors.lastname: Case
mesh_terms: D000328:Adult; D000368:Aged; D002469:Cell Separation / Q000379:methods; D002470:Cell Survival; D004996:Ethidium; D005260:Female; D005452:Fluoresceins; D005883:Gingival Crevicular Fluid / Q000166:cytology* / Q000276:immunology; D006801:Humans; D007958:Leukocyte Count / Q000379:methods; D007962:Leukocytes; D008297:Male; D008875:Middle Aged; D009504:Neutrophils; D010514:Periodontal Pocket / Q000276:immunology*; D010518:Periodontitis / Q000276:immunology*; D012044:Regression Analysis; D007507:Therapeutic Irrigation / Q000295:instrumentation / Q000379:methods*; D014343:Trypan Blue
chemical_list: D005452:Fluoresceins; D004996:Ethidium; D014343:Trypan Blue; C018506:diacetylfluorescein