Indirect Methods

Most of these studies - and especially those conducted on particulates - are limited to the exposure of cells to test chemicals that are either solubilized or suspended in the culture medium [64-71].Although this exposure condition may be adequate for soluble test materials, this may not follow the in-vivo exposure pattern of airborne aerosols, particularly for insoluble aerosols, due to unexpected alternation of their compositions and particle-medium or particle-cell interactions [72]. Such techniques of exposure may also ignore size, which is crucial in toxicity testing of inhaled particles.

Some research groups have employed sampling aerosols by filtration techniques, followed by an investigation of the effects of suspended and extracted particles, including those conducted on atmospheric aerosols [73-77] or cigarette smoke condensates [78-81]. The cytotoxicity of roadside airborne particulates was studied in rodent and human lung fibroblasts using the filtration technique [74]. Cytotoxicity was investigated using cell proliferation, (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS) and lactate dehydrogenase (LDH) in-vitro assays. Airborne particulates were sampled on glass-fiber filters using a high-volume sampler, after which the filters were sonicated using benzene-ethanol solvents; a crude extract was then obtained by evaporating the solvents to dryness. This extract was further fractionated by acid-base partitioning, and all extracts were dissolved in dimethyl sulfoxide (DMSO) for cytotoxicity assays [74].

Although filtration offers an advantage for the on-site toxicity assessment of aerosols, the technique usually requires sample preparation (e.g., extraction) in order to isolate the components of interest from a sample matrix. Ultimately, solubilization or suspension in the culture medium will potentially increase experimental errors and further toxicity interactions.

An indirect exposure technique has been developed in which samples of airborne formaldehyde were collected in serum-free culture media using an impingement method [82]. Cytotoxicity was investigated after treating human lung-derived cells with collected air samples. The objective of this study was to develop an in-vitro sampling and exposure technique that could be used for toxicity testing of soluble airborne contaminants, but with the potential for on-site applications. An average of 96.8% was calculated for the collection efficiency of airborne formaldehyde in serum-free culture media, which signified the potential application of this method for sampling airborne formaldehyde or other soluble airborne contaminants. The use of serum-free culture media as a collection solution for soluble airborne contaminants proved to be a simple technique, with no specific sample preparation or extraction steps required; moreover, this prevented any potential toxic interactions of the test chemical with other toxic organic solvents during preparation.

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