Alexa fluor 488 donkey anti chicken secondary antibody

To label Schwann cell nuclei, after treatment with ronidazole for 2 days Tg(gSAIzGFFD37A; UAS:NTR-RFP; UAS:EGFP) or Tg(gSAIzGFFD37A; UAS:NTR-RFP; zCrestHSP70:GFP) animals were fixed for 1 hour at room temperature in sweet fix (4% paraformaldehyde with 125mM sucrose in PBS) plus 0.1% Triton X-100 (Fisher, BP151). Animals were washed in phosphate buffer and incubated overnight at 4°C in primary chicken anti-GFP antibody (1:2000, Aves labs, GFP-1010) in incubation buffer (2 mg/mL BSA, 0.5% Triton X-100, 1% NGS). After washing in phosphate buffer, animals were incubated overnight at 4°C in Alexa Fluor 488 donkey anti-chicken secondary antibody (1:1000, Jackson ImmunoResearch, 703–545-155) in incubation buffer. After washing with phosphate buffer saline, larvae were incubated overnight at 4°C in Vectashield plus with DAPI (Vector Laboratories, H2000). Animals were mounted in agarose in a glass-bottomed dish and imaged in 1.5 μm slices using a x40 water immersion lens on a Zeiss LSM880 confocal microscope using Zen software. These samples were used to count total Schwann cell numbers.