The invention discloses an ultrahigh sensitivity
RNA in-situ detection method. The method comprises the following steps: S1, obtaining a tissue slice through a frozen section; s2, fixing is conducted for 15 min through 4% PFA; s3, 2x SSC rinsing is carried out twice, and each rinsing is carried out for 5 min; s4, rinsing with 70%
alcohol; s5, soaking in 70%
alcohol for 2 hours; s6, carrying out # imgabs0 # treatment for 15 minutes; s7, PBST rinsing is carried out once; s8,
protease K treatment is performed for 10 min; s9, rinsing twice with 4% BSA (
Bovine Serum Albumin); s10, 2x SSC rinsing is carried out twice; s11, sealing is conducted for 30 min through sealing liquid; s12, performing probe hybridization for 2 hours; s13, cleaning with a cleaning solution twice, 0.5 h each time; s14, incubation is carried out for 0.5 h through stepdavidin-hrp (stem davidin-human recombinant
protein); s15, 4% BSA cleaning is carried out five times, and each time is 5 min; s16, carrying out a TSA reaction for 15 min; s17, sheet sealing is carried out; and S18, observing and recording the
signal through a
microscope. The probe is divided into the primary probe and the secondary probe, the primary probe and the secondary probe are mixed to form the complete probe, a plurality of probes are designed for each
RNA molecule,
signal amplification is carried out, and
signal amplification is carried out through combination of
streptavidin coupled with HRP and
biotin of the secondary probe and through a Tyramide
signal amplification technology, so that signal detection with ultrahigh sensitivity can be achieved.