Boost Precision, Reproducibility for Critical LC Methods
Chromatographers who learned their craft twenty years ago may not have been familiar with autosamplers then, but today nearly every high performance liquid chromatography (HPLC) instrument includes an autosampler as standard equipment.
“Autosamplers existed in the early days of HPLC but many scientists preferred manual injection,” notes Simon Robinson, HPLC product manager at Shimadzu Scientific Instruments (Columbia, MD). Back when sample was plentiful, manual injection was a convenient way to flush the injection loop between runs, an operation most easily accomplished (at the time) by hand.
But the main reason most lab workers preferred manual injection was that early autosamplers were not very reliable. Robinson recalls horror stories where analysts set up a tray of twenty labile samples or so, then returned the next day to find the instrument stuck on sample number two and all the remaining samples “wrecked.”
Today 95 percent or more of HPLC systems from major manufacturers ship with autosamplers, a testament to the improved reliability and reproducibility of autosampler hardware and controls. “There’s been so much engineering focused on improving these instruments that it’s safe to say autosamplers are 100 percent reliable.”
Automation reduces errors
By automatically drawing from and injecting a predetermined set of samples, autosamplers spare lab analysts from uninteresting, repetitive work. Most autosamplers handle multiple sample containers, including microtiter plates, by default.
Even more time and drudgery can be saved when autosamplers are coupled with robotic sample preparation. But autosamplers’ major contributions to HPLC are reproducibility, repeatability, precision, and accuracy in delivering precise injection volumes. “An autosampler can do a much better job than any human working with a manual injector, particularly with small volume injections,” Robinson says.
Sample scarcity is one of the hallmarks of modern analytical science. Whether one works with biological samples, in forensics, pharmaceuticals, or food science, HPLC is called on to do more with less—to detect trace constituents or contaminants, more often than not within tiny samples. HPLC artifacts are common with manual injection, and many of these anomalies may be traced back to sampling and injection errors.
Autosamplers have eliminated these persistent sources of error, allowing analysts to target other possible sources of chromatographic anomalies. Now, analysts need only set up a tray of samples and make sure the correct sample is in the right vial. One would, therefore, be hard-pressed to find anyone involved in serious research, development, or analytic support using manual injection.
“For these reasons autosamplers make sense even for low-volume situations,” says Helmut Schulenberg-Schell, Ph.D., worldwide LC marketing manager for Agilent Technologies (Waldbronn, Germany), as well as for walk-up instruments shared by users and groups.
But are autosamplers for everyone?
So who would not routinely use an autosampler? According to Robinson, academic labs still rely heavily on manual sampling and injection because their primary objective, aside from data quality, is cost effectiveness.
Another area where autosamplers are often impractical, observes Schulenberg-Schell, is in preparative HPLC. Prep work tends to be specialized, one-off, nonroutine, and does not require the sensitivity or accuracy of analytical HPLC.
Autosamplers provide a level of automation, throughput, and productivity that is multiplicative. Using an autosampler, one chromatographer can operate multiple HPLC systems simultaneously, injecting many hundreds of samples after spending just a few minutes with the control software. This frees workers to perform other tasks like making mobile phases or analyzing data.
A systems approach to speed
Speed, throughput, and reduced sample volumes are critical for most analytical laboratories, and autosamplers play prominently among the various “fast” techniques adopted toward achieving those goals. Autosamplers have become critical enablers—like columns, pumps, and detectors—of these trends, and in doing their part they have required substantial re-engineering for both stand-alone performance and integration. As such, autosamplers have been a primary target of engineering efforts to shorten cycle times vs. simply run times. After all, an HPLC or UPLC system does not add much in the way of throughput or speed in reducing analysis time if the slowest link in the cycle still requires several minutes to re-equilibrate.
The wider adoption of UHPLC underscores the “need for speed” and the role of autosamplers within that scheme. “Now, suddenly,” says Schulenberg-Schell, “you can run ten times as many samples on an HPLC instrument, and the question of automation in sampling, injection, sample prep, and sample tracking becomes more urgent.”