Because adding PCR-enhancing agents can increase the yield of
the desired PCR product or decrease production of undesired
products (Chakrabarti and Schutt, 2001; Henegariu et al., 1997;
Binet and Maurelli, 2007; Kreader, 1996), we looked at whether
using glycerol, Dimethyl sulfoxide (DMSO) or Bovine Serum Albu-
min (BSA) in the multiplex PCR assay instead of water would
significantly improve the PCR efficiency or specificity (Fig. 5).While
the addition of 10% glycerol improved PCR specificity, it also caused
preferential amplification of ipaH over the other two targets. The
addition of 5% DMSO had no effect on the undesired product, and
the addition of 0.8 mg/ml BSA had a positive effect on some samples
and a negative effect on others (Fig. 5). The same trend was
observed with the 1 h and 1 h 40 min amplification runs (data not shown); addition of DMSO, glycerol or BSA caused the preferential
amplification of ipaH but did not improve the sensitivity of the
assay (data not shown).