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Looking for advice: Stable Isotopes

Grant RW Humphries, March 24, 2013
 

Hey everyone,

I'm looking for some quick advice on collecting stable isotopes in the field.

I am collecting blood and feathers from Sooty shearwaters, and am wondering:

1) For blood: Do I have to use a centrifuge to spin to blood after it is collected in the field? Or will it be okay to just preserve with Ethanol?

And:

2) If blood is collected, what is the timeframe I am looking at? I.e. does blood correspond to 1-2 weeks of feeding? 3-4 days? Or does it depend on whether or not I am using plasma vs. whole blood?

Thanks!! grant

Comments ( 7 )

Sjurdur Hammer

Sjurdur Hammer

  1. I had the blood, but the main reason was not due to the isotope, but because I wanted to analyse plasma for POPs. I'm not sure, but I think you can do SI analysis on whole blood, but others will be better to advice. If you need to centrifuge, you can potentially bring a portable hand-centrifuge (although I've only heard from unsuccessful attempts at that) or keep the samples on ice and shake them as little as possible. This would by my experience give you a couple of hours before the blood clots.

You should definitely not use ethanol if you're doing SI analysis!

  1. I'm not sure with regards to a differentiation between plasma and whole blood. As far as I understand blood (i.e. usually plasma) reflects a couple of days timescale.

James Grecian would be much better to advice. I will ask and get back to you.

Sorry for the half answers, and good luck!

Bill Henry

Best methods for preservation are freezing or drying (air, oven, freeze-dry) - (Barrow et al. 2008).

Yes, turnover rate for: plasma ~2-3 days, whole blood 1-2 weeks.

Breeding stage can correlate with si values.

If you want plasma and centrifuge - you will need a greater sample volume. Keep samples cool prior to centrifuging.

Collecting feathers? If not please do - 2-3 breast (outside brood patch) and subsample primaries.

Katharine Goodenough

Katharine Goodenough

Hi Grant,

For whole blood- best way is to air dry. I place a few blood drops on slide and let the slides air dry. If field conditions are a bit hectic, you can always use an Eppendorf to collect sample and then scrape it out. Freezing is always an option if you are using whole blood.

For plasma- I use lithium heparin as a preservative and then spin down the samples and pour off the plasma. Just keep in mind that any preservative will possibly alter carbon values. If you were to use this method, you could get information on both short term (plasma) and longer term (RBCs).

Turnover rates can be very species specific, but generally plasma provides dietary info within1-2 weeks, and whole blood is 4-6 weeks. The tissue turnover rate can depend a lot on whether the birds are in a state of growth or nutritional stress. Sears et al. 2007 had a nice paper on this.

Lisa Sztukowski

Hi Grant, You can use ethanol if air-drying, freezing, etc. are not feasible. But collect a base sample from each batch of ethanol you use as the SI signature changes by company and batch. I'm sure using heparin would be similar. Anything you add to the blood can change the signatures, but additives may be necessary to preserve the sample. I'm using ethanol for my samples as air-drying samples can grow mold in damp conditions.

Turnover rates can vary as Katharine Goodenough mentioned. I don't have access to the Sears et al 2007 paper at the moment (if someone could send it to me I would appreciated it). But from what I've read serum (plasma if you use heparin) provides information on assimilated protein over the previous 4-5 days, whereas red blood cells correspond to the prey consumed in the previous 4-5 weeks (Hobson and Clark 1992a, b, 1993).

Based on how my samples congeal with ethanol, you should centrifuge the samples prior to adding ethanol if you want both serum/plasma and red blood cells.

Hope all it going well in the field.

Steffen Oppel

Steffen Oppel

Hi Grant,

it all very much depends on what exactly you are trying to find out. Centrifuging blood to separate RBC and plasma components is valuable if you suspect that the birds may have been foraging on isotopically distinct diets over the last week (plasma) vs. the last 1-2 months (RBC). If there is no reason to believe that they have, then you can use whole blood.

Storing blood in ethanol may affect the carbon isotope signature, but whether that is of significance again depends on your question and your study system. In remote areas where freezing isn't an option storage in ethanol can work ok - you just need to be careful when you interpret the values.

Useful papers (the Barrow paper mentioned above is on turtles I think): Bugoni, L., McGill, R. A. R. & Furness, R. W. 2008. Effects of preservation methods on stable isotope signatures in bird tissues. Rapid Communications in Mass Spectrometry, 22: 2457-2462.

Hobson, K. A., Gibbs, H. L. & Gloutney, M. L. 1997. Preservation of blood and tissue samples for stable-carbon and stable-nitrogen isotope analysis. Canadian Journal of Zoology, 75: 1720-1723.

If you use whole blood, then the isotope signature will usually be dominated by the RBC component, and thus reflect what the birds were foraging on over the past 1-2 months. However, the turnover is only relevant if the birds shifted between isotopically distinct food sources - which you may expect for birds that just returned from migration to the North Pacific, but may not be the case for birds that stay in the Southern Ocean for months before you catch them.

cheers, steffen

Grant RW Humphries

Grant RW Humphries

Awesome! Thanks a lot everyone. That's a big help. And thanks for responding so quickly!

Cheers G

Rocio Moreno

Rocio Moreno

Hi Grant,

Personally, if you can avoid it, I wouldn't use ethanol at all. If you just can freeze the samples, it would be great. That way, if you have some blood/plasma left (you will!because you need a very small amount for SIA), you can use it for something else. Last year I used priceless whole blood from 12 years ago that was in the freezer, fortunately, without ethanol!

Cheers,

Rocio