AQUIOS STEM: Workflow comparison between the AQUIOS STEM System and the FC500 Stem-Kit

Authors: Andreas Böhmler, Maja Diemel

Affiliation: Beckman Coulter Germany, Krefeld

On this page you will:

• Learn about the differences between the AQUIOS STEM System and the FC500 Stem-Kit
• Discover the benefit of the AQUIOS STEM System

Objective

Today more than 90,000 hematopoietic stem and progenitor cell (HPC) transplantations are performed annually worldwide for the treatment of several hematological malignancies as well as for nonhematological indications.

Clinical laboratories rely on commercially available IVD solutions for CD34+ HPC enumeration in order to avoid time- and resource-consuming validation of user-defined tests. Most reagent kits and software packages were developed as a response to the 1996 and 1998 ISHAGE Guidelines, which have not been updated to meet the evolving demands of diagnostic laboratories, especially in terms of automation capability. In hemato-oncological laboratories, HPC samples often arrive as emergency (STAT) samples in the laboratory and require immediate attention, disrupting the routine workflow. Any issues with these samples duplicate efforts and thus increase the potential for human error. For these laboratories, it would be desirable to integrate HPC samples into the normal workflow, ideally in a way that minimizes the risk for sampling mistakes or other issues, as the analysis of CD34+ HPC is time critical.

The present case study compares the workflow of the newly developed AQUIOS STEM System for automated CD34+ enumeration against its predicate method, the Stem-Kit for the FC500 flow cytometer (both Beckman Coulter, Inc.), in terms of turnaround time, hands-on time, and number of potentially error-prone steps that may affect specimen or result integrity.

Overview of traditional flow cytometry workflows

Many traditional flow cytometry solutions for CD34+ enumeration, such as the Beckman Coulter Stem-Kit for FC500, require manual preparation of samples, manual creation of worklists, manual data review, and manual tabulation of numerical data. This workflow can result in longer run times, more hands-on time, and requires more experienced operators.

In order to perform a CD34+ enumeration test with the predicate method (Stem-Kit on FC500), the operator is required to manually check for instrument alignment, prepare controls to verify and adjust fluorochrome spillover (compensation), prepare and run process controls, and finally, prepare and run the actual patient/donor samples. Resulting data needs to be verified and manually transferred to the laboratory information system (LIS). Reagent and quality control (QC) logbooks are typically kept manually. See Figure 1 for a representative manual flow cytometry workflow.

Figure 1. Representative manual flow cytometry workflow example

Overview of the AQUIOS CL Flow Cytometer Workflow

The AQUIOS CL is an automated system that performs most of the preparatory steps leading to a test result, so that most manual preparation steps are eliminated.

For the AQUIOS STEM System for CD34+ enumeration on AQUIOS, the operator needs to load the reagents on the system once, start up the instrument, run Flow Check and QC samples, and is then ready to load and analyze patient samples throughout the remaining workday (Figure 2).

Figure 2. Representative AQUIOS CL Flow Cytometry System workflow example

Comparison Protocol

The purpose of this case study is to evaluate the AQUIOS CL Flow Cytometer design for CD34+ enumeration, and to model its resultant workflow through comparison with the Beckman Coulter Stem-Kit on FC500 (predicate method). This case study estimates the following workflow parameters:

• Turnaround time – For alternative systems, the time duration begins with the first sample preparation step and ends with completed test results. For the AQUIOS CL system, time duration begins with the sample placed in the autoloader and ends with the completed test result for the last sample being displayed. For both systems, the time does not include QC.
• Operator hands-on time – The time required by the user to physically perform the steps listed in the provided test procedures according to the manufacturer’s Instructions for Use.

Note: Time points were captured by videotaping sample preparation and analysis processes and modelled using an in-house workflow analysis software.

Sample processing turnaround time and operator hands-on time

This test case consisted of scenarios with either 1 sample or a batch of 10 samples of mobilized peripheral blood, prepared in duplicate plus negative control. The AQUIOS STEM System was compared directly to its predicate method, the Stem-Kit on FC500. Data are shown for tests that consist of duplicate runs plus negative control.

For 1 specimen sample, time to result from sample preparation to result generation did not show a major difference (56:19 minutes for the predicate method vs. 53:54 minutes for AQUIOS; Figure 3a, left), as the process steps are mainly determined by the incubation times with the antibodies and red blood cell lysis reagent.

However, operator hands-on time could be reduced by 95%, from 6:23 minutes for the predicate vs. 0:20 minutes on AQUIOS (Figure 3a, right).

For a batch of 10 specimen samples (an average daily workload in a mid-sized stem cell transplantation center), these differences become more prominent. Time to result from specimen preparation to result generation went down from 2:30:04 hours for the predicate method to 1:56:54 hours for AQUIOS (Figure 3b, left), with operator hands-on time being reduced from 55:44 minutes for the predicate method vs. 3:20 minutes for the AQUIOS STEM System (Figure 3b, right).

As also for a manual process, certain steps can be performed in parallel (pipetting the next sample during the incubation time of the first sample, etc.), the overall time for 10 samples is shorter than the sum of 10 times one sample.

Figure 3. Sample processing turnaround time and operator hands-on time for one CD34+ sample (a) or a batch of 10 CD34+ samples (b) on the FC500 with Stem-Kit (Alternative Method, grey bars) and AQUIOS STEM System (AQUIOS, red bars).

Assuming 10 samples per workday over the period of one year, the AQUIOS STEM System saves a lab hundreds of hours of valuable tech time each year. In addition, the faster time to result when running an average workload of 10 samples per day supports time-critical assays such as CD34+ enumeration.

Process time and operator hands-on time for Quality Control (QC) procedures

In a regulated environment, system performance needs to be controlled with appropriate quality and process controls for both the instrument settings as well as the test to be run. For both the Stem-Kit on FC500 and AQUIOS, daily verification of the flow cytometer's optical alignment and fluidics system is performed with Flow-Check Fluorospheres, an assayed suspension of fluorospheres (fluorescent microspheres). As process controls for the verification of the analytical parameters of interest, the predicate method uses lyophilized CD34+ cells that are added to a normal blood sample (Stem-Trol Cells), while the AQUIOS STEM System uses liquid preparations of stabilized human leukocytes (lymphocytes, monocytes and granulocytes) and erythrocytes that have lysing, light scatter, antigen expression and antibody staining properties representative of those found in human whole blood specimens (AQUIOS STEM CD34 Control Cells).

The total time needed for QC procedures related to CD34+ enumeration is reduced from 2:02:22 hours for the predicate method to 1:09:44 hours on the AQUIOS system. Of note, QC needs to be completed before patient/donor samples can be run, which leads to a time savings of 52:38 minutes per day (Figure 4a, left). For a 5-day work week, this sums up to approximately 4.5 hours only for the QC procedures (Figure 4b, left).

Operator hands-on time for QC procedures is reduced from 12:22 minutes (predicate) to 1:51 minutes (AQUIOS) per day (Figure 4a, right), resulting in a total hands-on time savings for QC processes of 52 minutes per week (Figure 4b, right).

Figure 4. Process time and operator hands-on time for Quality Control (QC) procedures on the FC500 with Stem-Kit (Alternative Method, grey bars) and AQUIOS STEM System (AQUIOS, red bars) per workday (a) and a 5-day work week (b).

Summary

In all test case scenarios, the AQUIOS STEM System required substantially less hands-on time than the predicate method. For a typical 5-day work week with 10 specimen samples per day, the AQUIOS STEM System reduces manual workload by approximately 6 hours, allowing the lab to allocate resources more efficiently. In addition, the AQUIOS STEM System reduces the overall turnaround time from sample preparation to patient result, which is essential for a time-critical test such as CD34+ enumeration

The AQUIOS STEM System is a quantitative automated solution that performs the enumeration of CD34+ hematopoietic stem and progenitor cells in a “no-wash” sample preparation process. Since this system is intended to be an automated analyzer with hands-off processing of samples from specimen introduction to results reporting, it is referred to as a Load & Go flow cytometer. The automation features differentiate the AQUIOS STEM System from alternative methods (including its predicate method, i.e., FC500 with Stem-Kit), where many process steps need to be performed manually. The all-in-one approach of the AQUIOS STEM System is intended to assist in time savings and to help increase the workflow efficiency of a modern laboratory.