A pre-hybridisation treatment with different SDS concentrations significantly affects the WMISH signal.
L. stagnalis larvae of different ages were subjected to pre-hybridisation treatments with varying amounts of SDS and then hybridised with anti-sense probes to beta tubulin
(J-O) and COE
(P-U). For all genes and larval ages, treatment with 0.1% SDS did not generate consistent or strong WMISH signals (A, D, G, J, M, P and S). Treatments with both 0.5% and 1% SDS produced strong WMISH signals for beta tubulin and engrailed in larvae aged three to five days post first cleavage (dpfc), with high spatial resolution (inlet in K). For COE 0.5% SDS outperformed the 1% SDS treatment (T vs. U). Black stars indicate optimal treatments. Note that some treatments produced equally good results. The most consistent results (defined as constantly good signals among genes and ontogenetic stages with little variation between individuals within an experiment) were achieved with 0.5% SDS (examples shown in B’, E’, H’, K’, N’, Q’ and T’). Larvae in A-C and M-R are shown from an apical perspective, larvae in D-F are viewed ventrally, G-I laterally and J-L and S-U dorsally.