Ph. In 'T Veld et al., The certification of a reference material for the evaluation of methods for the enumeration of Bacillus cereus, J APPL MICR, 86(2), 1999, pp. 266-274
A reference material containing Bacillus cereus was certified by the Commun
ity Bureau of Reference (BCR) for its number of colony-forming particles (c
fp) in 0.1 ml reconstituted capsule solution. To this end, a batch of appro
ximately 15000 capsules was produced and tested for its homogeneity and sta
bility. The variation in the number of cfp between capsules (homogeneity) w
as not found to be significantly different from a Poisson distribution. Sta
bility was tested for extended periods at storage temperature (-20 degrees
C) and at various higher temperatures up to 37 degrees C for 4 weeks to sim
ulate transport conditions. Only at 37 degrees C did a small but significan
t decrease in the number of cfp occur. At -20 degrees C, no decrease in the
number of cfp was observed over a period of about 4 years. For certificati
on, 12 laboratories determined the number of cfp on two agars: Mannitol Egg
-Yolk Polymyxin agar (MEYP, incubated at 30 degrees C) and Polymyxin pyruva
te Egg-yolk Bromothymol blue Agar (PEMBA, incubated at 37 degrees C). The c
ertified geometric mean value on MEYP after 24 h of incubation was 53.4 cfp
0.1 ml(-1) of the reconstituted capsule solution (95% confidence interval
51.7-55.2) and on PEMBA, 55.0 (95% confidence interval 52.8-57.4). Based on
these certified values, user tables were constructed specifying the 95% co
nfidence limits when testing a smaller number of capsules, as would be done
in individual laboratories. Based on the information on homogeneity, stabi
lity and the certification study, the BCR decided to certify the material a
s CRM 528.