The most recent recommended procedures for STED sample preparation can be found in the Leica technical notes below. The documents are relevant for systems with 592 nm, 660 nm and 775 nm depletion lasers. The UCL Super-resolution Facility currently only has a 592 nm depletion laser, so not all dye combinations will be possible. Please email the facility staff if you are unsure what dyes to use. Also, the UCL system does not currently have 3D STED capability. In other words, it cannot improve resolution in the axial dimensionZ dimension, only in X and Y.
| Anchor | ||||
|---|---|---|---|---|
|
Quick Guide to STED Sample Preparation by Wernher Fouquet
STED 3X Sample Preparation by Louise Bertrand and Sarah Crowe (hosted by Yale University)
...
The technical notes above contain detailed information about dyes but essentially , if you are imaging a single channel , Leica recommends using Oregon Green; but , but Alexa 488 also works well. If you want to image a second channel you will have to use a long Stokes shift dye. Leica recommends Abberior Star440SX or BD Horizon V500. Star 440SX is available conjugated to secondary antibodies but you will most likely need to use the streptavidin conjugated form V500 as it is designed for use in flow cytometers and is only available directly conjugated to antibodies against leukocyte antigens. This means your secondary will have to be conjugated to biotin.
...