Further, the correlation between caries experience and extracellu

Further, the correlation between caries experience and extracellular levels of heat shock protein-70 cause (eHSP70) was studied in saliva samples of these students. MATERIALS AND METHODS The present study comprised 147 subjects. All these cases were selected from female students aged 16�C21 and attending undergraduate classes at local colleges in Punjab, India. Students were randomly selected from a list of candidates in UG classes, and every individual had an equal chance of selection. A simple probability random sampling method was used in obtaining information about caries experience (DMFT) and calculus and their relationship to eHSP levels in saliva samples. The researchers adhered to WHO guidelines while preparing the questionnaire for data collection.

Ethical approval of the study was obtained from college authorities, and written consent was obtained from the students. Oral examination of all subjects was performed by a trained dentist (RK) who examined all exposed and accessible surfaces for caries. The students were examined while seated in an ordinary chair, in broad daylight and facing away from direct sunlight. The dental examination for carious teeth involved sequential assessment of teeth, from 1 to 32. The dental examination was performed with the help of illumination devices, and data were recorded for analysis. A mirror and explorer were also used during the examination. Compressed air was used to dry teeth and to remove debris to allow better visual examination. The instruments used were sterilized after every single use.

The site presenting dental caries (decayed teeth, DT) at a cavity level was recorded, and filled (filled teeth, FT) and missing teeth (MT) were counted separately; data are presented as DMFT, DT, FT, and MT. Subjects were asked about the level of caries and calculus as well as the duration of symptoms in a questionnaire. The questionnaire, given to the sample group, also investigated oral hygiene, lifestyle, and snack habits of the subjects. ELISA for eHSP assay The primary antibodies used for ELISA were monoclonal anti-HSP70 (clone-BRM-22, SIGMA), which recognizes both constitutive and inducible forms of HSP70. The secondary antibody used was anti-mouse IgG, peroxidase linked antibody (Bangalore Genei). All other chemicals and reagents were the purest available commercially from local suppliers.

Saliva sample collection Saliva was collected during the forenoon between 10.00 and 12.00. Participants were told to first use mouthwash and then allowed to chew on a cotton ball. When stimulated saliva accumulated in their mouth for 4�C5 min, it was transferred to a collecting vessel. Saliva samples were precleared by centrifugation (15,000 rpm, 4��C for 15 min), and sterile filtered with GSK-3 0.22 pore size filters and stored at ?20��C for further use. Saliva HSP level was determined by indirect ELISA. Saliva was diluted to a 1:1 ratio with a coating buffer (Na2CO3-NaHCO3 buffer 0.05 M, 9.6).

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