Work has been carried out in an initiative to establish a U.K.-based measurement infrastructure for aerosol and bioaerosols. The initiative is supported by the National Measurement System Policy Unit (NMSPU) of the U.K. Government Department of Trade and Industry (DTI) under the Valid Analytical Measurement System (VAM) scheme. The overall aim of the work is to examine the appropriateness and practicality of validating and certifying microbiological reference materials for bioaerosol standards. One of the prime uses of such bioaerosol standards is the characterisation of samplers and assay methods. Candidate microbiological materials were selected from a survey of mainly U.K. industrial (and others) user needs. The microorganisms selected are Escherichia coli (National Collection of Industrial and Marine Bacteria, NCIMB 86), Saccharomyces cerevisiae (National Collection of Yeast Cultures, NCYC 1337) and Penicillium expansum spores (Rothamsted isolate no. C2636). Culturing techniques are often used in conventional aerobiology to enumerate the numbers of microorganisms. This publication reports on the programme of work to examine the effects that aerosolisation parameters and residence time in the spray and collection liquids can have on bioaerosol culturability. Data showed that suspensions of S. cerevisiae (NCYC 1337) cells can be maintained at an average culturable fraction of 0.7 for a period of 3 h in a suitable buffer solution maintained at the ambient room conditions prevailing in the studies (20 degrees C and 30% RH). This suspension can be aerosolised to produce a bioaerosol with a culturable fraction that depends on spray suspension age and collected aerosol age. The culturable fraction of this bioaerosol was found to decrease with spray suspension age and with collected aerosol age. This strain of the species may not be suitable as a microbiological reference material. The results of the work with E. coli (NCIMB 86) showed that aerosolisation reduced the culturable fraction in the bioaerosol to close to zero, although the culturable fraction could be maintained at approximately 0.24 (stationary phase) and 0.33 (log-phase) for cells maintained in the spray suspension for up to 3 h. It is concluded that this strain of E. coli (although defined to be a fairly robust strain of this species)is not suitable as a standard bioaerosol or microbiological reference material. The results of the studies with P. expansum (Rothamsted isolate No. C2636) spores showed that aerosolisation had little general effect on the culturability of the bioaerosol (average culturable fraction of 0.25). This observation is to be expected with fairly robust microorganisms such as fungal spores. In view of their robustness, P. expansum, spores are recommended as a microbiological reference material, and that a standardised aerosol of this test microorganism should be prepared according to the methodology described in this paper for culturing, preparing the spray suspension, aerosolisation under the correct ambient conditions, collection and assay. The results of the tests indicate that traditional culturing techniques underestimate cell/spore numbers in the collected bioaerosols of the microorganisms used. Total count techniques are therefore recommended for the determination of cell/spore numbers in collected aerosols of the microorganisms used in the tests. The culturability of P. expansum spores is unaffected by aerosolisation, and total number may be determined either by total counts or by culture counting taking into account the initial culturable fraction in the spray suspension. It should be noted that different methods for counting total cell numbers may give different results.