Locate the vial labeled "0.5 mL neat Triton X-100". Make a stock solution of 100 ppm (0.01% v/v) Triton in methanol by adding 10 µL of Triton X-100 to 100 mL of analytical grade methanol and mixing thoroughly. This stock solution will serve as an EIA negative control and will be used for adding Triton X-100 to extracts prior to solvent exchange and for bringing the Triton X-100 concentration of sample and/or standard EIA tubes to the target level of 10 ppm (final concentration) during EIA analysis.

The following is a generic extract preparation procedure which is appropriate for samples which have gone through the full cleanup procedure prior to GC/MS analysis (using immunoassay compatible internal standards).

For other samples, consult the appropriate Application Note . Use of this EIA for screening analysis depends upon the same amount of each sample being introduced into the EIA. This quantity (the matrix load in "mg sample equivalent per EIA tube") is dictated by the target sensitivity and must be determined in advance.

Consult Table 3 to determine the appropriate matrix load.

Table 3: TEQ Levels (ppb) at Which Screening Decisions are Made:
(based on three standards and six different matrix loads).

How to use Table 3:
Locate your desired decision level (in ppb TEQ) in the body of the table (the orange cells), then read the "mg Sample Equivalent per EIA tube" at the top of that column. This is the amount of sample you must use per EIA tube. If your decision level occurs in two table columns, use the left one (corresponding to the lower standard in the left column). This will allow less sample to be used, giving less potential for matrix interferences. The comparison of your sample to the appropriate standard in the left column allows you to make the decision you want, as illustrated by the examples below the table.

Example 1: When using 1 mg sample equivalent per EIA tube, a screening decision at 10 ppb would be made by comparing the sample to Standard 2 (1 ppb 2,3,7,8-TCDD).

Example 2: To make a 10 ppt screening decision by comparison of the sample to Standard 2, a matrix load of 1000 mg sample equivalent per EIA tube would be required.

Example 3: When using 10 mg sample equivalent per EIA tube, a screening decision at 10 ppb would be made by comparing the sample to Standard 4 (10 ppb 2,3,7,8-TCDD).

It is best to exchange only enough sample for immediate analysis, plus the minimum amount to cover waste in pipetting. Leave the rest of the sample in toluene or other less volatile EIA-incompatible solvent.

Redissolving the sample in 30 µL of methanol will allow recovery of one or two 10 µL aliquots for single or duplicate EIA analysis.

Use a clean glass tube or conical vial for evaporating each sample to be tested. Add an appropriate amount of Triton X-100 in methanol solution to each evaporation tube or vial.

If the sample will be redissolved in 30 µL of methanol, then add 30 µL of 100 ppm Triton in methanol.

Evaporate solvent completely under dry gas stream. The original solvent must be completely gone. Recovery will not be adversely affected if the sample remains under the gas stream for a few minutes after reaching dryness.

Complete removal of 100 µL toluene generally takes at least 15 minutes at room temperature. Larger starting volumes may require gentle heat and more time to complete the evaporation. During solvent exchange the Triton X-100 functions as a "keeper", similar to the way dodecane or tetradecane are often used.

When the original solvent is completely evaporated, the PCDD/Fs stay with the Triton, not on the glass, so they are easily redissolved in methanol.

Add methanol to each tube and mix vigorously for 15 to 30 seconds to redissolve the detergent and sample residue. Sonication or longer mixing times have generally not been found to be necessary.

Samples should contain 100 ppm of Triton X-100 when redissolved in methanol. Immediately pipet the redissolved sample into the EIA tube which has been prepared (10 µL per EIA tube as noted in Section J5 below).

This minimizes the evaporative loss of methanol. Add methanol to redissolve the next sample only when you are finished adding the previous sample to the prepared EIA tubes.

If larger reconstitution volumes are used, samples redissolved in methanol can be stored for later analysis if desired.