Introducing Our System Solution for High Throughput Quality Control - HTQC 

Let’s start with a question: What does high throughput mean?

For us in the lab, high throughput usually means analyzing dozens, hundreds, or even thousands of samples per day in the laboratory or on a particular instrument. But how can this be achieved and what is necessary to perform fast, reliable and automated measurements? In this blog we take a closer look at those requirements and how they are implemented in our HTQC system.

Speed

High throughput HPLC applications most often require fast methods. This could refer to both the overall method runtime and the cycle time of the injection module.

Fast gradient applications up to 1200 bar can easily be realized with the HTQC system thanks to its two included UHPLC high pressure gradient pumps.

Fig. 1–2  HPG pumps of HTQC system, left: P 8.1L (max. pressure 1240 bar), right: P 6.1L (max. pressure 1000 bar)

We will not go into the details of switching from classical HPLC to fast, high-resolution UHPLC methods here, but the introduction of UHPLC methods can significantly speed up the process. 

Let's do the math for a hypothetical application. Imagine Giorgia (just a randomly chosen name 😊) needs to perform quality control on a pharmaceutical sample in 1998. Obviously, she must have travelled through time, as she was too young to work in a lab back then, but you get the point. Using a classical HPLC method, she needs around 75 minutes for each sample, including equilibration, analysis, and cleaning the 250 x 4.6 mm ID column by backflushing. This is a typical method used in QC labs all over the world. For an eight-hour working day, Giorgia can analyze around 6 samples a day.

Transferring the same method to UHPLC gives Giorgia a significant productivity boost. With analysis time, equilibration time and backflushing time all reduced to less than 6 minutes per sample, she can now analyze around 90 samples in a single working day. 

Why are two gradient UHPLC pumps necessary in our HTQC system, you might ask yourself? We will give you an explanation for this a bit later.

Let’s continue to think about how to improve the overall speed. The cycle time of the injector can sometimes be very time-consuming due to washing steps and the injection procedure itself. That’s where our liquid handler LH 8.1 enters the stage.

Fig. 3  KNAUER LH 8.1 Liquid Handler

With its clever feature to use overlapped injections you can effectively save time because the injection for the next run can be already prepared while the measurement is still in progress, without compromising necessary cleaning steps of the injection system.

Fig. 4   Schematic buildup of injections with and without overlapped injections

Let's do the math again. Giorgia is very happy that she can use overlapped injections in her QC analysis. She now saves about 1 minute per sample and can thus analyze around 105 samples per day.

Sample Throughput

Another crucial point for high throughput analysis is the number of samples your system can handle or store. Again, the LH 8.1 shows its benefits due to the opportunity to use up to three robotic coolers when a version with a longer rail is chosen. Three coolers, each with three drawers, a total of nine 10x13 sample trays: 1170 vials! And they are temperature controlled as well.

Fig. 5–6 Robotic Cooler with two 10x6 trays, LH 8.1 (887 version) with two Robotic Coolers  

Combined with the typical low eluent use of UHPLC, this setup gives us the possibility to extend our working hours from 8 to 24. Let's see what this means for Giorgia and her QC lab. The same UHPLC method, including the function for overlapping injections, can now be used around the clock. This allows around 320 samples to be processed per day. Do you think this is enough, or is there any possibility of speeding up this method even further? Watch our blog title carefully: We just can´t get enough! 😉

Automation

Besides the common 6 port-2 position injection valve, our HTQC system is equipped with an additional valve. This special column switching and backflushing valve allows you to operate two alternating columns in a single UHPLC system. While one column is running the analysis, at the same time the second column can be backflushed and equilibrated in reverse flow direction. This is a major advantage for reducing matrix effects due to complex sample matrices typically found also in QC labs. Furthermore, this contributes to the overall runtime of the method.

 

Fig. 7 Exemplary flow scheme of special column switching valve

Parallel Processing

Do you remember what you asked yourself at the beginning? Why are they using two UHPLC gradient pumps? If you’ve read carefully so far, then you have already discovered a clue in the previous section. The second gradient pump is necessary for backflushing one column while the other one is switched into the analytical stream by our special valve.  

Fig. 8 Schematic illustration of column cleaning, equilibration and analysis with special backflush valve

Giorgia immediately ordered a second UHPLC pump and the special valve, and tested them in her QC lab. She now has a complete HTQC system and is obviously very happy.

Why? Let's do the math one last time: Thanks to all the features and automation, Giorgia can now clean one column while the other runs a sample. She can analyse an incredible 600 samples per day! Compared to the HPLC method she used in 1998, the number of samples processed per day has increased from six to 600, an increase by a factor of 100!

Summary

We started with a question and now let´s end with one as well: What have we done to increase the throughput? 

First, we did a scale down from a classical HPLC to a fast UHPLC method. Then, we added a second pump and a special column switching and backflushing valve to our UHPLC system. Combined with the liquid handler LH 8.1 and its outstanding sample storing capabilities, the HTQC system is a versatile configuration for high throughput analysis.

Needless to say, this system can be coupled to almost any detector that can be used in HPLC. Due to the possibility of very fast operation, mass spectrometric methods are a perfect fit!

Fig. 9 Complete flow scheme of HTQC system with highlighted system components

For further information on this topic, please contact our authors:  kramer@knauer.net, m.myrach@knauer.net