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Slowfade gold mounting medium with dapi

Manufactured by Thermo Fisher Scientific

SlowFade Gold mounting medium with DAPI is a reagent designed for use in fluorescence microscopy applications. It is formulated to reduce photobleaching of fluorescent dyes, including DAPI, which is a nuclear counterstain. The product is intended to be used as part of the sample preparation process for microscopy analysis.

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3 protocols using slowfade gold mounting medium with dapi

1

Drosophila Larval Tissue Staining

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Drosophila larva transitions from larval stage L1 to larval stage L3 by intermittent molting. Prior to pupariation, L3 larvae stop feeding and migrate to a pupariation site on the side of a vial. Wandering L3 larvae were bisected and inverted in PBS, and all tissues were fixed in 4% formaldehyde/PBS for 30 min, permeabelized in 0.2% Triton X-100/PBS for 4 hours, and blocked in 3% BSA/PBS for 1 hour. All antibody stainings were performed in 3% BSA/PBS, incubation of primary and secondary antibodies were O/N. PBS was used for all rinses and washes (3× each for primary and secondary antibody incubation steps). Antibodies used were as follows: Chicken anti-RFP 1:2,000 (Rockland, 600-901-379); Goat anti-GFP 1:3,000 (Rockland, 600-101-215); Donkey anti-Goat Alexa Fluor 488 (Life Technologies, A11055); Donkey anti-Chicken Cy3 (Jackson ImmunoResearch, 703-165-155); and Donkey anti-Mouse Alexa Fluor 594 (Life Technologies, A21203). All secondary antibodies were used at 1:500. Tissues were dissected off the cuticle and were mounted in SlowFade Gold mounting medium with DAPI (Life Technologies, S36938). See Kockel et al. (2016) (link) for a detailed protocol.
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2

Immunostaining of Drosophila Larval Neurons

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L3 larvae from cross of StanExnovel insertion with a line harboring a LexA operator- GFP reporter transgene (LexAop2-CD8::GFP; Pfeiffer et al., 2010 (link)) were dissected in PBS and fixed in 4% Formaldehyde/PBS for 30 min, permeabilized in 0.2% Triton X-100/PBS for 4 hr, and blocked in 3% BSA Fraction V/PBS for 1 hr. Incubation of primary and secondary antibodies were O/N in 3%BSA/PBS at 4° Celsius using a platform rocker. All specimens were rinsed (1 min) and washed (20 min) three times with PBS after antibody incubations. Primary Antibody: Goat anti-GFP 1:3000 (Rockland 600-101-215). Secondary antibody: Donkey anti-Goat Alexa488 (Life Technologies, A11055). All samples were mounted in SlowFade Gold mounting medium with DAPI (Life Technologies, S36938).
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3

Multicolor Immunofluorescence Staining

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All tissues were fixed in 4% formaldehyde/PBS for 30 min, permeabilized in 0.2% Triton X-100/PBS for 4 hr, and blocked in 3% BSA/PBS for 1 hr. All antibody stainings were performed in 3% BSA/PBS, incubation of primary and secondary antibodies were overnight at 4°. PBS was used for all rinses and washes (3× each for primary and secondary antibody incubation steps). Antibodies used: Chicken anti-RFP 1:2000 (Rockland, 600-901-379). Goat anti-GFP 1:3000 (Rockland 600-101-215). Mouse anti-Tubulin 1:5000 (Sigma T5168). Donkey anti-Goat Alexa488 (Life Technologies, A11055). Donkey anti-Chicken Cy3 (Jackson ImmunoResearch 703-165-155). Donkey anti-Mouse Alexa594 (Life Technologies A21203). All secondary antibodies were used at 1:500. All samples were mounted in SlowFade Gold mounting medium with DAPI (Life Technologies, S36938).
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