OK so you followed our recent series on the Carry-Over Effect problem that can lead to inaccurate Blood Alcohol Content results in the posts:
- A large problem in Gas Chromatography: No uniform standard for GC run position or composition
- The Carry-over Effect: Lack of Blanks between tests leads to false positive or inflated BAC results
- Carryover effect part Deux: Autodilution may be part of the problem for false blood results in DUI
But the person that comes from the Government lab to testify at trial, your under-trained, non-technically educated, non-PhD and non-analytical chemist, smiles and says:
"Ahhhh Ha! Attorney (fill in the blank with your name), the internal standard that we use will catch all"
This is not unlike the "deus ex machina" endings to the old time movies and even some modern ones like James Bond.
Ever notice how the bad guy captures James Bond and rather than killing him thinks up some sort of massively contrived mechanism that 007 can always seemly escape from based upon massively unlikely events. It leaves you saying "Ah come on now". Doesn’t it?
You see a "deus ex machina" ending is a plot device first really developed and classified in ancient Greek theater. It is generally frowned upon because, much like the James Bond movies, it leaves the watcher, the reader or listener saying "Ahhhhh, come on now." It’s cheap.
The "deus ex machina" plot device occurs when an outside force completely resolves a previously intractable problem suddenly and abruptly. It is solved with a contrived and artificial introduction of a new character, ability, or object. It is anti-climatic typically.
So, don’t let the State witness succeed by introducing a contrived and artificial new argument.
This post will hep de-mystify the "internal standard dues ex machina retort to the carry-over argument"
First, what the Government will say about the use of the internal standard:
An internal standard is a method whereby a known and constant amount of a traceable and pure analyte is added into the various samples in the gas chromatography run. In simpler terms, the samples are spiked in a known and fixed/constant amount adding a different chemical. The key to selecting the internal standard is that it must be similar in nature, but not likely endogenous/native, meaning present, to the unknown samples to be analyzed later. What this means is that in the case of ETOH examination of unknowns, the near universal consensus is to use the chemical n-propanol to spike the samples. N-propanol is close to ETOH, but all well setup gas chromatographs should be able to separate out these compounds with ease (R>1.5).
So n-propanol is added to the within run calibrators, positive controls, negative controls and the blanks.
This creates within each headspace vial a known amount of something. If that known amount remains constant from vial to vial and from vials to the ones that created the calibration curve, you should be able to compare the ratio of the known to the unknown as established in the calibration curve to get the quantification of the analyte of interest, in our case ETOH.
Signal from the analyte that we are looking for (ETOH) is compared with signal from the internal standard (the introduced and spiked n-propanol) to find out how much analyte of interest, in our case ETOH, is present.
As long as the concentration of standard is known AND REMAINS CONSTANT, the correct concentration of analyte of interest (ETOH) can be derived. If you violate this principle of the need for a constant, precise and known amount of internal standard across the run and with the calibration curve, then the whole system falls apart.
The internal standard is delivered by volumetric pipetting.
If the internal standard is not exactly introduced remaining constant and precise from sample to sample, the result is inaccurate. The ratio means nothing. The reported quantification of the analyte of interest (ETOH) is in doubt.
The violation of that most essential principle inherent in the "internal standard dues ex machina retort to the carry-over argument", that need for the internal standard to be introduced in a constant, precise and known amount can happen rather easily even by the most experienced and well intentioned analysts.
As mentioned above the internal standard is delivered by volumetric pipetting. This is the weak link.
This is particularly possible where an automated system is not used. A good and accredited lab will keep pipetting tolerance certificates when they order and get new pipettes from a manufacturer. A good and accredited lab must also have pipetting proficiency tests for each analyst. This is why a good criminal defense and DUI attorney should always ask for both the manufacturer’s certificate of the pipetting tolerance and the particular analyst’s pipetting proficiency results.
(Above: Proper protocol for "conditioning" a pipette)
As one can see, this is an involved process.
But what about the automated pipetting system? It’s after all automatic, right? Right, but…..
This even occurs for autodiluters as the pippettes may not be pre-wetted or "conditioned".
-Justin J. McShane, Esquire, Pennsylvania DUI Attorney
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