After washing, reaction with streptavidin and Alexa Fluor 488 (green) conjugate (1∶150; Molecular Probes, Eugene, OR, USA) was conducted at room temperature for 1 h.
After extensive washing, reaction products were quantified by Cerenkov counting.
After extensive washing, reaction was revealed with 3,3'-diaminobenzidine hydrochloride (Sigma, St . Louis MI) and 0.02% peroxide hydrogen, followed by counterstaining with Mayer's hematoxylin.
We attempted to recover immunoreactivity in lichen extract-treated samples by filtering reactions through a 10 12 kDa molecular weight cutoff (MWCO) membrane and washing reactions with 50 volumes of phosphate buffered saline (PBS).
cDNA synthesis, labelling, hybridization and washing reactions were performed according to the NimbleGen Arrays User's Guide (V 3.2).
After washing, reactions were developed with peroxidase color substrate (KPL, Gaithersburg, MD), and quenched by the addition of 1N H2SO4.
The cDNA synthesis, labeling, hybridization and washing reactions were performed according to the NimbleGen Arrays User's Guide (V 3.2).
Blocking, addition of secondary antibody and washing reactions were performed according to the manufacturer's instructions for the BondMax staining system with Bond Polymer Refine Detection Reagents (Leica Microsystems, Wetzlar, Germany).
After a final wash, the reaction was developed by the addition of orthophenyldiamine (OPD) to each well.
Tissue sections were cut at 40 µm and washed in cold phosphate-buffered saline, followed by washes with reaction buffer.
