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Super-resolution microscopy allows biologists to go beyond the 200 nm diffraction limit of traditional confocal microscopes. Through the use of innovative optical techniques, super-resolution microscopy offers exciting new insights into the intracellular world.
Stimulated emission depletion (STED) microscopy reduces the size of the point spread faction (PSF) through the use of a pair of lasers. The first laser works just like a regular config excitation laser pulse, exciting the fluorophores and causing it to fluoresce. That first laser pulse is quickly followed by the STED beam: a donut-shaped pulse of a laser at a longer wavelength. The ring of fluorophores excited by the STED beam is instantly reverted back to a ground state, effectively bleaching them. The result is only a tiny collection of fluorophores sitting at in the hole of the donut STED that are able to fluoresce, reducing the PSF and taking resolution beyond the diffraction limit. With STED microscopy, you can achieve a lateral resolution of around 20 nm lateral, and an axial resolution of 40–50 nm.
To achieve super resolution, it's important to select an antibody conjugate that has excellent emission crossover with the STED laser wavelength to achieve the required level of saturation. Leica, for example, recommends the ATTO 647N and ATTO 655 dyes for use with the Leica TCS STED microscope. Antibodies should also ideally be preabsorbed to minimize background from non-specific binding.
Our ATTO dye-conjugated secondary antibodies are all preabsorbed to IgGs of multiple species and offer you
Our STED range includes
Immunocytochemistry/immunofluorescence - Goat Anti-Mouse IgG H&L (ATTO 655) preadsorbed - STED Microscopy (ab245991). HeLa (human epithelial cell line from cervix adenocarcinoma) cells were stained for alpha Tubulin using a mouse Anti-alpha Tubulin antibody at 1/500 dilution, followed by ab245991 at 1/1000 dilution in ICC/IF analysis (red).
Immunocytochemistry/immunofluorescence - Goat Anti-Rabbit IgG H&L (ATTO 647N) preadsorbed - STED Microscopy (ab245992). HeLa (human epithelial cell line from cervix adenocarcinoma) cells were stained for histone H3 trimethyl Lys27 using a rabbit polyclonal Anti-Histone H3 trimethyl Lys27 antibody at 1:500 dilution, followed by ab245992 at 1/1000 dilution in ICC/IF analysis (red). Nucleolin is stained green.
Immunocytochemistry/immunofluorescence - Goat Anti-Rabbit IgG (ATTO 655) H&L (ab245993) preabsorbed - STED Microscopy. HeLa (human epithelial cell line from cervix adenocarcinoma) cells stained with Histone H3 trimethyl Lys27 rabbit polyclonal antibody at 1/500 dilution and ab245993 at 1/1000 dilution in ICC/IF. Green: Nucleolin mouse monoclonal antibody.