Modifications to a Standard Buccal Collection Protocol: Effects on Human DNA Yield

A standard mouthwash protocol (a single 10-mL swish of mouthwash for 45 sec) was modified in an attempt to increase the amount of human buccal cell DNA per collection and to reduce the percentage of low yielding human DNA collections (<4 μg). A group of 22 healthy individuals donated a buccal sample each week for several weeks according to the standard protocol without or with one of the following modifications: (1) decreasing the volume of mouthwash, (2) having participants externally rub or not rub their cheeks before donating a specimen, (3) donating two consecutive specimens at each collection, (4) substituting saline for mouthwash, and (5) having individuals expectorate into mouthwash. There was no significant difference in the amount of human DNA collected when 10 mL or 5 mL of mouthwash was used. Externally rubbing cheeks before donating did not significantly alter the amount of human DNA collected, regardless of whether it was one or two donations. Addition of a second donation resulted in 24% to 50% more human DNA than from only a single donation, regardless of whether 5 mL or 10 mL of mouthwash was used, with or without cheek rubbing. With two donations, the percentage of low-yielding human DNA samples was reduced up to 31%. Substituting saline for mouthwash resulted in a significantly lower amount of human buccal DNA collected, regardless of cheek rubbing, and a higher number of low-yielding samples. Expectorating directly into mouthwash while externally rubbing cheeks performed the best (17.70 g human DNA), followed by swishing 2–10 mL of mouthwash before expectorating (12.26 μg). Both protocols had 95% of their samples yielding at least 4 μg of human DNA without increasing cost; however, when selecting a collection protocol the age and health status of the cohort needs to be considered.