Mushroom exposure
Measurement of airborne spore for determination of
b-D-glucan level was made by drawing air through a
110-mm diameter glass fiber of 0.6 mm porosity
(GB-100R, Toyo Roshi, Tokyo, Japan) at a flow rate of
500 l min 1 for 30min using a high volume air sampler.
The air inlet of the sampler was attached at the subject's
nose level. The filters were shaken for 10 min in 10 ml
distilled water. Twenty-five microlitres of the extract
and 100 ml glucan specific lysate (Fungitic G test;
Seikagaku Kogyo, Japan) were added. The plate was
incubated in a spectrometer, and the kinetics of the
ensuing color reaction was read photometrically.
Sampling consisting three measurements involved three
filters in the cultivating room, three in the harvesting
and packing area, and three in the ofice room. The
limitation of detection was 20 pgml 1 for b-D-glucan.
The mean concentrations of airborne b-D-glucan were
1 ngm3 in the cultivating room, 3 ngm 3 in the
harvesting and packing area and 3 ngm 3 in the ofice
room.
Mushroom spore antigens preparation
The spores of Bunashimeji were collected from the
mushroom in a sterilized method at this farm, and
cultured in Sabouraud's glucose broth. The cultured
spores and their proteins were extracted with 50%
ammonium sulfate. The sample was centrifuged at
3000 rpmfor15min and the pelletwas dialysed with dis-
tilledwater.
Serum precipitins to Bunashimeji
The double di¡usion test for precipitating antibodies
for HM was performed according to the method of
Ouchterlony (13). Fifty microlitres of antigen and/or
serum were added to opposite well of 1% agarose plates
and monitored for 72 h at room temperature. The fol-
lowing antigens: Micropolyspora faeni, Thermoactinomyces
vulgaris (prepared in our laboratory) and Aspergillus fumi-
gatus (Hollister-Stier, Spokane, Washington, U.S.A.)
were also tested in the same manner.