Figure 2. An example of the capabilities of the P450RGS cell-based assay for increasing site characterization.

ERDC TN-DOER-C19

December 2000

or DRE) on the DNA. The binding of the complex to the DNA triggers the transcription of several

related genes (known as a gene battery), which code for the synthesis of a series of messenger RNA

(mRNA), which in turn are translated into proteins. The P450RGS cell line has been modified by the

insertion of plasmid DNA that links the DRE region to a transgene, the firefly luciferase gene. Bind-

ing of the TCDD-receptor complex to the DRE results in the production of the enzyme luciferase,

which can then be quantified in a luminescence assay. The gene insertion allows for an extremely

sensitive endpoint, luminescence, compared with the fluorescent endpoint typically measured for

the most commonly used native TCDD gene battery product, ethoxyresorufin-O-deethylase

(EROD). Although this is an example of TCDD, the general model of specific receptor interaction

leading to transcription of specific gene batteries is similar for a wide variety of chemical

contaminants.

The P450RGS assay illustrates the potential savings and benefits offered by cell-based assays. Al-

though gas chromatography-mass spectroscopy (GC-MS) is the definitive method for identifying

and determining the precise amount of dioxin and dioxin-like compounds present in an environmen-

tal sample, the method is expensive (~$1,200 -

$2,000 per sample). P450RGS is far less expen-

sive at about $200 per sample, and its use can result

in major cost savings when used to screen multiple

sediment cores to determine which samples should

be confirmed by GC-MS.

A recent field test of the cell-based assay demon-

strated the benefits of increased site characteriza-

tion (McFarland et al., in preparation). In this

study, five cores were taken from each of 13 sites

in a channel requiring dredging (Figure 2). Each

core was sectioned into 3-ft (0.9-m) intervals to

provide depth profiles for TCDD contamination

(Figure 3). All samples were analyzed in triplicate

100

with the P450RGS assay. At five of the sites, five

TEQ

to ten additional cores were taken and composited

Range

for GC-MS analysis. Due to the high cost of

TEQ predicted by cell assay

TEQ from chemistry data

GC-MS, no replicates were analyzed. The in-

creased coverage provided by the P450RGS assay

Figure 2. An example of the capabilities of the

(155 samples analyzed in triplicate for cell-based

P450RGS cell-based assay for

assays versus five samples with no replication for

increasing site characterization. The

GC-MS) resulted in a detailed and statistically ro-

correlation coefficient for the chemical

bust characterization of the contamination bound-

analysis and cell-based assay data

was r2 = 0.71. The TCDD Equivalent

aries within the channel. The P450RGS results

(TEQ) from chemical data represents

shown in Figure 3 indicated that the bulk of the

one composite sample, while the

contamination at sites 8, 12, and 13 was in the top

TEQ from the cell assay represents a

layer of the material to be removed. If only the

composite of 15 to 60 individual

GC-MS data were used to characterize the site,

extracts (depending on the depth of

most of the dredged material would be considered

the core)

contaminated. However, obtaining information of