Fluorescence Spectra Graphs
20190223 - I would prefer to have updated an earlier Pulse article, but linkedin/Microsoft appear incapable to enabling this. Earlier articles related to this are at:
https://www.dhirubhai.net/pulse/resolution-blues-meets-21plex-salute-fluorescence-basic-mcnamara/
https://www.dhirubhai.net/pulse/18plex-flow-cytometry-from-brilliants-when-catch-up-george-mcnamara/
Here are several spectra graphs I generated with https://www.biolegend.com/spectraanalyzer
Alexa Fluor 6plex (more are available, BioLegend has these in the Spectra Analyzer).
8 QDots (ThermoFisher) and 2 eFluor NC (nanocrystals = QDots, ThermoFisher/eBiosciences [acquired by ThermoFisher]).
5 Super Bright (ThermoFisher) + 4 of the BV Brilliant Violets (BD Sirigen, also available from BioLegend).
7 BV Brilliant Violets (BV750 not shown, since overlaps emission of Zombie NIR) + Zombie NIR (BioLegend DNA counterstain -- I recommend this instead of wasting 405nm excitation -> blue-cyan-green-yellow on DAPI or Hoechst 33258 or 33342).
6 BUV Brilliant UltraViolets (BD Sirigen) + Zombie Red (BioLegend, DNA counterstain).
8 (and a graph with 5) prototype NIRvana Sciences chlorins, bacteriochlroins, etc. ... NIRvana's scientific founder, Jonathan S. Lindsey (NCSU) has published extensively on these (see Chem Rev. and PhotoChemCAD 3.0). Lindsey lab and NIRvana are not the only chemists working on chlorins -- nature has been using -- for billions of years -- related chlorophylls to turn sunlight into energy and biomass.
Above: Bruce Pitner linkedin profile background - NIRvana Sciences 8plex prototype emission spectra ... https://www.dhirubhai.net/in/bruce-p-75a7b23/
Above: Bruce Pitner 2016 ISAC CYTO poster - NIRvana Sciences 58plex prototype emission spectra vs BV QDots emission spectra widths.
Below: Absorption spectra (excitation spectra are usually, not always, very similar) ... all the fluorophores excite well with 405 nm laser, common on laser scanning confocal microscopes -- or 395 nm LED excitation now (becoming) common on widefield microscopes (as Mercury arc lamps, Mercury "metal halide" lamps, and Xenon arc lamps are retired).
I would like to see (and try in our lab at JHU) the tandems of:
BUV395 --> NIRvana 8plex
BV421 --> NIRvana 8plex
BB510 (or BB515) ==> NIRvana 8plex.
Adamas Nanotechnologies "blue" fluorescent nanodiamonds (FND) --> NIRvana 8plex.
I suggest the Adamas FNDs (they have a 2019 paper with 8plex from blue to NIR) should also REFLECT (aka scatter) light well, thanks to very high refractive index (R.I. 2.6 for diamonds, so same for nanodiamonds), for example, 140 nm FNDs, with 405 nm lase light (or 355 nm laser or LED on instruments BUV capable), would be sub-resolution, and likely label (i.e. Ab-FND, or better nanobody-FND).