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In multiple ISH reactions, cross controls (mix of antisense and sense cRNA probes followed by detection using the double/triple detection protocols) were carried out for all probe combinations and detection protocols and the results were compared with single ISH for the antisense cRNA probes using the appropriate detection protocol for these probes.
In ISH control labelings, substitution of antisense cRNA probes by an equivalent amount of hapten-labeled sense cRNA probe, omission of cRNA probes and cross controls (mix of one antisense cRNA probe of interest with another sense cRNA probe and detection using the double or triple detection protocols) lacked unspecific staining.
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The upper layer of the specimens (WMA or hot mix asphalt control mix) was mixed and compacted at different temperatures in order to simulate different application conditions.
Waste is automatically transfered to the sealed inner reactor, where the computer controls mixing, air flow, temperature, and moisture.
As negative controls, mixes without cDNA were used.
As controls, mixed bone marrow chimeras made with Ighb CD1d +/+ and Igha CD1d +/+ bone marrow cells were prepared.
Setup of the PCR assays included positive and negative controls, mixing experiments to rule out bias, and repeat experiments to assess reproducibility.
Spiking primers were mixed to create a pan-positive control mix for the assay (Additional file 4: Table S4).
Comparative studies on control mix against trial mix were carried out.
A total of four mixes, including the control mix, were designed.
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