Suggestions for Field Sampling

Sample preparation guidelines for C, N and S stable isotope analysis prior to shipment. (Updated 3/12/06, Boston University Stable Isotope Laboratory)


Our minimum sample sizes are normally 1 µmol C, 2 µmol N and 20 µmol S (12 ug C, 28 ug N and 640 ug S). If possible, please adjust your sampling to meet these minimum requirements; it would be better to have several times these amounts in case problems arise in the analysis.

If you cannot meet these minimum sample sizes, we will need to make special arrangements. Our ultimate limits for measurement are about 0.5 µmol N (7 ug N), and 0.2 µmol C (2.4 ug C), although samples this small usually have significant blank corrections.

Before sending off samples, they should have been dried at 60 µC in glass vessels or on aluminum foil, or in paper bags (large leaf or soil samples). Avoid plastic drying containers because they can partially melt if your temperature control on the oven is not precise, and melted plastic can especially contaminate d13C measurements. If samples are larger than a few milligrams, they should also have been ground to a fine powder (0.50 grains of powder per mg) before shipping. Use a Wiley mill (#40 mesh) or a mortar and pestle for grinding; liguid nitrogen will help in grinding the material in a mortar and pestle. Once dry and powdered, samples can be stored at room temperature in glass or plastic vials.


For wood, leaves, roots, large algae, etc., collect 1-5 grams. Dry at 60ºC, grind to a fine powder in a Wiley Mill (#40 mesh) or with a mortar and pestle.

For phytoplankton and periphyton loosened from hard surfaces, filter water through a precombusted (500ºC, 4 hr) glass fiber filter (GF/C). Keep filtering until filter clogs. If that is not practical, try to filter enough water to discolor filter. If convenient, record amount of water filtered, and chlorophyll concentration. Dry filters at 60ºC. Send a blank filter for all sample sets.


Collect at least 5 mg. For small animals less than 1 gm, use the whole animal, but remove gut contents by dissection or by holding the animals alive until gut contents are cleared. You may have to pool many individual small animals to meet the 5 mg minimum (e.g. zooplankton, nematodes, etc.). Smaller samples can be done, but you must first contact the lab manager. For larger animals, use a scrap of muscle tissue from a carcass, or liver tissue. Dry at 60ºC, and grind to a fine powder with mortar and pestle.

Sediments and Soils 

Collect 5-15 gm samples, sieve to remove large rocks, shells, etc. Dry at 60ºC. Grind to a fine powder in a Wiley Mill or with mortar and pestle.

Dissolved Inorganic Carbon (DIC)

Collect 250 ml or more of water in a glass bottle, add 1 ml of 1% HgCl solution, stopper and seal with electrical tape. Store in a cool place. Do not filter water sample, as this will result in the loss of dissolved CO2.

Dissolved N (nitrate and ammonium) 

Sample size is often a problem in these analyses, but by modifying our Fisons elemental analyzer coupled to the Finnigan Delta-S IRMS, we are able to analyze samples down to ca. 1 uM. If you are also interested in ammonium, acidify the water with concentrated sulfuric acid to a pH 2 to prevent volatile losses of N. Please collect 500 to 1000 ml in acid-washed polyethylene bottles. Acid conditions will inhibit microbial use of N. 

If DON is a problem, we have worked out an alternate procedure for DIN analysis. Please contact the lab for further information. The procedure involves collecting 500-1000 ml water, filtering the sample, then acidifying to a pH of 2 with concentrated sulfuric acid.

Special Notes for Sulfur Samples

Although we do not run sulfur samples, we are including this section for investigators who may decide to collect and send their samples to other labs.

Due to the low sulfur content of wood, d34S cannot usually be measured. For other plant and animal samples, you may be interested in organic sulfur rather than total sulfur that includes inorganic sulfate. Inorganic sulfate can be a large contaminant, especially in marine or coastal samples, but can be removed by repeated leaching of the samples with distilled water.

Procedure: resuspend a ground sample in distilled water, shake for 5 minutes, centrifuge, discard supernatant, repeat. Redry and grind sample. If you want C and N measurements on the same sample, send a separate bottle with unleached sample. Some soluble organics are lost during leaching and may affect d13C and d15N.

Dissolved Sulfur (sulfate and sulfide)

Sulfates. Sulfates should be precipitated with barium chloride and sent as a white salt (BaSO4). Many lake samples need to be concentrated by boiling to 100 ml before precipitating sulfate, since sulfate does not precipitate readily when µmol/l. Precipitation procedure: bring sample to boiling in a Pyrex beaker, add concentrated HCl until the pH <2. Add 1 ml of prefiltered 10% barium chloride solution. Let sample stand for 20 minutes at a low boil, then cool (covered) overnight. Next day, filter precipitate onto a Whatman 42 or Nucleopore 0.2 µpore size filter. Fold, place in a glass or plastic scintillation vial, and dry at 60ºC. If no precipitate forms, raise the pH to 5 or 6 with NaOH additions, and repeat procedure. If still no precipitate, check sulfate concentration--it is probably too low.

Sulfides dissolved in water. Free sulfides should be precipitated in a 2% zinc acetate solution (final concentration). Let solution stand overnight at room temperature, siphon off supernatant. The next steps are designed to remove carbon contamination (acetate) from the sample. Transfer white flocculant zinc sulfides to centrifuge tube, spin down, discard supernatant, wash pellet by resuspending in distilled water, centrifuge again, discard supernatant; then flush centrifuge tube with nitrogen gas, seal tightly and send tube.

Solid phase sulfides. Sulfides in minerals and sediments are often separated from surrounding rock or organic matrix by chromium reduction and acid distillation. In these procedures, sulfides are converted to H2S and trapped in a 2% zinc acetate solution. If recoveries are near 100% in the distillation and trapping, the zinc sulfides can be used for d34S measurements. Treat the zinc sulfides from traps as described above for sulfides dissolved in water. 

Sulfides can also be trapped as cadmium sulfide, but should be boiled 5-10 minutes with 1% silver nitrate solution to convert them to silver sulfides. Concentrate silver sulfides by filtration and scrape from filter. Dry at 60ºC.


Put samples in glass or plastic vials or in aluminum foil (small animals). Label distinctly, provide accompanying data sheet with sample number, description, and your name and telephone number. If known please give exact or approximate C, N and S content of samples (e.g. %C, µmol C/g, µmol N/l, etc.). Also please state if samples contain any carbonates (shells, CaCO3 precipitates), as we need to remove carbonates before d13C analysis of organic fractions. A short description of your project and goals would be helpful, but is not necessary. It makes sample preparation more interesting for the students in the lab if they know something about the project.

Send package to :

Robert Michener Boston University Stable Isotope Laboratory Boston University Department of Biology 5 Cummington St. Boston, MA 02215 (voice) 617-353-6980 (fax) 617-353-6340 (email)

For further information: