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20 μl is an excessive volume for mounting 22×22 mm coverslips, so the top coverslip should be easily removed by flicking the slide. If the top coverslip does not readily slide off the fiber coverslip, place the microscope slide in a container with enough volume of 2×SSC (after hybridization) or PBS-T (after an amplification layer) to cover the slide and gently shake the slide until the top coverslip floats off.
