I'm preparing an experiment for this summer and I want to dye bivalve post-larvae (4mm long). I will use 150mg/L of calcein with 1 hour incubation of juveniles. Do you know how many times the staining will be efficient? Can I use directly sea water to mix the calcein? Or do I have to use miliQ water and sodium bicarbonate?
Thanks
The papers cited below should help you out. I personally use Alazarin Red to dye post-larvae (2-3mm). Usually just mix the dye in the seawater that the clams are kept in for ~1 week at a 100ppm concentration - stains them nicely and makes them easy to distinguish.
With respect to calcein, as far as I know it is not necessary to use sodium bicarbonate, but doing so enhances the solubility of the calcein (see Wilson et al. 1987; citation below).
Moran, AL and Marko, PB. 2005. A simple technique for physical marking of larvae of marine bivalves. Journal of Shellfish Research 24: 567-571.
Riascos, J. et al. 2007. Suitability of three stains to mark shells of Concholepas concholepas (Gastropoda) and Mesodesma donacium (Bivalvia). Journal of Shellfish Research 26: 49-49.
Wilson, CA et al. 1987. Calcein as a fluorescent marker of otoliths of larval and juvenile fish. Transactions of the American Fisheries Society 116: 668–670.
Thank you Jeff !
I'll look at your suggestions soon, I think they will help me a lot.
Best regards.
These might also help:
1. Andresen, H., Dorresteijn, I., and van der Meer, J. (2013). Growth and size-dependent loss of newly settled bivalves in two distant regions of the Wadden Sea. Marine Ecology Progress Series 472, 141-154.
Matthijs van der Geest, Jan A. van Gils, Jaap van der Meer, Han Olff, Theunis Piersma, Suitability of calcein as an in situ growth marker in burrowing bivalves, Journal of Experimental Marine Biology and Ecology, Volume 399, Issue 1, 15 March 2011, Pages 1-7
Pierrick,
Your mail-adress isn't working. Please provide another? Mail with Pdf keeps bouncing back.
Pierrick,
we used Calcein on a number of adult and post-recuitment mussel species. Success-rate was reasonable and improved greatly if bivalves could be made to gape (open) while in the solution. This was most easily achieved by moving around the water (by pump of swirling by hand) around in the tank. Once you are sure that most of the bivalves have opened up for a short while, Calcein should be incorporated. We dissolved our Calcein in natural seawater as our marine bivalves do not like opening up in freshwater. Calcein does not dissolve easily, so we often had to heat the water until Calcein was dissolved and we then cooled the solution down and aerated it a bit before use. Hope this helps.
Sven,
okay for your method's description, it look easily realizable with good results.
In Thébault et al., 2006, they suggest to prepare a solution with Mili-Q water + sodium bicarbonate to dissolve calcein (acid) and have a good pH, near to pH 8 like sea water. I certainly will try this to kinds of methods.
Thanks for your help !
I think tha is conversation is interesting because I also work with calcein in pectinid, but not to do with the waste, anyone know if chlorine inactivates calcein? as scrap?, I must give some treatment to remove seawater? Thanks