The Christie Hospital in Manchester is one of Europe’s leading cancer centres, treating over 40,000 patients a year. The histopathology service provides diagnostic tumour pathology not only for the hospital’s own patients but also for those from other hospitals across the region and sometimes even further afield. The immunohistochemistry (IHC) service provided by the histopathology laboratory is extensive, with over 50,000 slides processed each year. This service is incredibly important and it is essential that the laboratory keeps up to date with developments in the field to provide a fast, efficient and reliable service. The laboratory therefore opted for the latest development in automated IHC systems – but more later.
The rest is history
Immunohistochemistry was first described in the 1940s using immunofluorescence for the detection of antigens in frozen sections. However, it wasn’t until the 1970s that the methods were applied successfully to routine formalin-fixed, paraffin wax-embedded (FFPE) tissue.
Two significant developments in the 1970s helped to shape the field of IHC. The first was the introduction of hybridomas to make monoclonal antibodies. These cell lines are capable of producing large quantities of primary antibodies that are made available commercially. The second development was the application of enzyme digestion to unmask antigens previously thought to be lost in FFPE tissue.
These two developments meant that there was a large increase in the number of antibodies produced for diagnostic purposes. Then, in the early 1990s, the use of heat-induced epitope retrieval (HIER) was first described. This led to an explosion in the number of commercially available antibodies as more antigens could be detected than previously thought. Much effort has also been applied to improving the sensitivity of the detection methods available, with advances from the direct method through to avidin-biotin systems and more recently to the sensitive micropolymer detection kits.
The major applications of IHC in diagnostic pathology have created a real need to standardise the procedure. Moreover, with the development of individualised cancer treatments based on the differential expression of prognostic markers, the requirement for standardisation is now paramount. The need for automation to standardise the technique has led to the introduction of numerous commercial systems. The first automated platforms appeared in the early 1990s and manufacturers are continually striving to develop systems designed to improve patient care.
Innovative automation
The automation currently available is generally robust and helps the laboratory to achieve a good quality of IHC staining and standardisation. Until recently, however, no system offered a large capacity and throughput while remaining easy and flexible to use. Neuropathology and techniques requiring the application of IHC on double width slides have also been overlooked until now.
Most histopathology laboratories can identify with the scenario of increasing workloads, a reduction in or deskilling of staff, recruitment difficulties and consolidation of services. This comes with the added pressures of becoming Lean, meeting targets to improve turnaround time, and reducing overall costs. In order to achieve this and improve service provision, there is a need for innovative products such as multiplexing and efficient, flexible automation. The intelliPATH FLX offers all this and much more.
The intelliPATH FLX mimics the manual IHC staining process on a flatbed system, truly automating the horizontal staining methods recognised as optimal for IHC staining. This instrument is not only speedy but has a large capacity. There are five slide racks, each with the capacity for up to 10 slides, accommodating up to 50 standard slides per run. This permits a significant throughput of up to 150 standard slides in a nine-hour working day and an additional 50 standard slides overnight. The increased slide capacity and throughput not only means quicker turnaround times but also the need for fewer IHC instruments in the laboratory. This frees precious space, means that less time is spent validating and maintaining equipment, and a reduction in overall costs.
Double width slide racks are also available for the intelliPATH FLX. This is a first for IHC automation. Currently, double width slides can only be processed manually, or the area of tissue to be stained taken from the large block and mounted on a standard slide. This leads to a delay in obtaining the result and involves an increased amount of staff input. The double width slide racks for intelliPATH FLX hold up to five slides. With a click of the mouse the operator is able to designate a slide rack for standard slides or double width slides. This means that both sizes can be accommodated during the same run. This feature will help departments that process large slides to become more Lean and reduce the turnaround time and the quantity of reagents used.
Special features
The intelliPATH FLX has many special features that help to make it fast and efficient. For example, it has a large-volume probe capable of aspirating up to 5 mL of reagent, which enables it to dispense on to 10 slides before it returns to collect more reagent. It also has a ‘no touch’ ultrasonic sensor that detects reagent level and sends a signal to the computer. This eliminates the need for time-consuming probe washing. Both these features, together with intelligent software scheduling and simultaneous X–Y motion, optimises the time to results.
Although the speed and capacity of the system is important to help laboratories meet those all-important targets, many departments require a system that is flexible and does not dictate how staff should work. The intelliPATH FLX offers flexibility in more ways than one. Whether your workflow involves running large slides batches, discrete small batches or even a combination of batches and short turnaround time (STAT) urgent slides, the intelliPATH FLX provides a solution, as it features multibatch processing (MBP) technology. This allows the operator to schedule the workload around the particular needs of the laboratory service, pathologists and patients.
The intelliPATH FLX platform provides true continuous random-access workflow by allowing the operator to add new batches of slides on demand. The system’s MBP technology enables the laboratory to run cases on a first-in, first-out basis, as each tray is run independently with separate start and completion times. This makes it easier to reduce turnaround-time and supports unprecedented levels of service. Furthermore, MBP enables the intelliPATH FLX to prioritise STAT samples. This ensures that urgent samples are processed fast, but the intelligent scheduling of the software ensures that batch delays are eliminated, as other runs will not be disrupted.
Onboard flexibility
Other features of the system that make it flexible include the onboard mixing and cold spot. The onboard mixing of enzymes and chromogens eliminates the time it takes for the operator to mix solutions. Using the refrigerated area (cold spot) on the staining platform, labile chromogens and enzymes can also be loaded and mixed onboard prior to application to the slide, eliminating the need for the operator to pause the run. This feature enables the user to ‘walk away’ or set up the intelliPATH FLX for overnight or delayed start runs, thereby increasing the laboratory’s throughput and efficiency.
In addition, the system permits user-defined protocols to be created with no restriction on the types of protocol that can be run simultaneously. It can be operated with multiple protocols (no limit), any number of incubation times (one minute to 24 hours) and multiple detection systems (no limit) simultaneously. The dual buffer inlets also allow simultaneous processing of protocols that require individual buffers.
The flexibility of the system ensures that it is optimised for multiplex (double) staining, which is becoming increasingly popular. The system provides a simultaneous testing platform for multiplex stains that demonstrate morphologically distinct markers using different chromogens, supplying superior diagnostic data while reducing slide workload.
Intuitive and easy to use
With so many special features you would expect the intelliPATH FLX to be quite complex to set up and run. However, the software is Windows-based and incredibly intuitive and easy to use. The 2D Matrix Mode ensures positive slide and reagent identification (ID), eliminating manual data entry and reducing sample ID errors. A 2D matrix scanner (barcode reader) is incorporated on all systems as a standard component.
The system is also supplied with two barcode label printers. Matrix (2D) labels can be printed for slides as well as for all reagents, which eliminates the need to purchase expensive barcoded reagent vials. However, use of the bidirectional LIS interface capabilities achieves even quicker results as duplicate data entry in both LIS and intelliPATH FLX software is eliminated.
Should your workload demand multiple intelliPATH FLX systems, it is possible to link up to four instruments to one PC, which permits sharing of valuable information for reagents, protocols and case lists. This allows lot number, expiry date and remaining reagent volume to be tracked, and reports generated regardless of on which system a patient case or protocol has been processed.
Feel the quality
Quality of results remains the most important factor when evaluating immunostainers. Assessment at evaluation sites and at sites using the intelliPATH FLX routinely demonstrates that results compare favourably with those obtained manually or using other automated platforms. As with any probe-based system, possible carryover of reagents can lead to false results if the washing of the probe is inadequate. However, the absence of carryover with the intelliPATH FLX demonstrates the effectiveness of the probe washing step. The wash step for the slides is also excellent, with only one 5-mL wash required between each reagent to guarantee clean staining, which helps to reduce run time and the need for repeats.
Internal calibration of the system at the start of each day only takes a few seconds. This helps to ensure accurate and reliable operation with no user intervention, reducing the amount of hands-on time required to support the system.
Maintaining good records is very important for quality control and accreditation but can be quite time-consuming to complete manually. All data entered in the intelliPATH FLX software and also information pertaining to each patient case and individual protocol and reagent validation is recorded in the extensive reports database. Graphs can be generated for easy comparison and all information can be exported an Excel file for data manipulation and statistical analysis.
A question of retrieval
Debate continues in IHC automation about onboard versus off-board retrieval. Onboard retrieval allows the operator to load slides and walk away, but certain aspects are often overlooked in relation to productivity and flexibility. To maintain the best productivity it is essential that smooth workflow with reduced bottlenecks is achieved. Performing HIER in a separate Antigen Access Unit (AAU) can increase throughput considerably. Large numbers of slides can be pretreated in as little as 30 minutes using a buffer that will simultaneously dewax and pretreat. The slides can then be loaded continuously on the intelliPATH FLX system during the course of the day. As the HIER step is not performed on the automated platform, run times can be reduced. Additional batches of slides can be treated while the system is in operation and will be ready to load as soon as one of the five slide racks becomes available.
The new AAU has been redesigned with a programmable walkaway capability, delayed start facility and a cooling fan for more rapid antigen retrieval. Furthermore, a USB port enables the laboratory to utilise a flash drive to track and save run data for quality control purposes. The pressure gauge has a new feature in which the needle remains at the highest pressure reading during the run, providing validation that antigen retrieval has occurred properly.
Ultimately, the AAU offers the operator a choice. It can pretreat up to 96 slides using up to four different buffers at any time and temperature in a single run, making it an excellent tool for both HIER and special stains. This combats the inflexibility of onboard retrieval, which often restricts buffer, time and temperature to factory-set protocols. Retrieval is a very important step that ensures excellent quality of the end result. Giving the operator control of this step using a standardised system and buffers ensures that quality results can be achieved for all antibodies.
The Christie Hospital experience
Two intelliPATH FLX systems were installed at The Christie Hospital in Manchester in November 2009. Laboratory staff members were slightly nervous about replacing the existing technology as it was capable of processing a large volume work each day. Karen Graham, laboratory manager, says: "We were worried that nothing would cope with the numbers we process. However, two IntelliPATHs cope comfortably with our workload, but we would have needed four or five of the 30-slide fully automated systems and don’t really have the space in our IHC laboratory. Also, because we have only had to install two intelliPATH systems, it has helped to keep costs to a minimum as we do not have to support and maintain multiple instruments.”
Karen continues: “Workflow is now much quicker through the laboratory since we have had the intelliPATH systems installed. Programming of the system is very simple and we can input all of our patient cases quickly at any point in time when they are received in the laboratory and print the labels for slides. This means we have to spend less time batching everything in order to write the slides and program the instruments, which helps to eliminate the backlog of slides we have experienced previously. In fact, the area that now causes the only bottleneck is in section cutting – we are physically unable to cut slides fast enough to keep up with the throughput of the intelliPATHs.”
The Christie Hospital is also implemented multiplex double stain antibodies and detection systems. The consultant histopathologists in the department have found the staining quality to be excellent and that the ability to view more information on one slide has proved very useful.
Biomedical scientists and consultants have been very happy with the quality of the results. Advanced practitioner Catherine Billington says: “It is always a worry when changing systems as much time and effort needs to be put into the re-titre of primary antibodies, and it can be a worry that there may be a dip in the quality of results. However, there has been a real improvement in the quality, and the results are even better than I was expecting. Also, the support received has been excellent. It was a bonus to know that Menarini support was always available at the end of the telephone, should it be needed, while getting to grips with the new system. The company provided a lot of support to re-titre our antibodies and this helped to alleviate some pressures from us so that we could maintain our service while switching systems.”
Advanced practitioner Liz Beckett adds: “The system is very easy to maintain, the automated cleaning cycle really takes care of itself and requires very little input from the operator. All staff members have found the system very simple to operate and have got to grips with the software very quickly. On a day-to-day basis, this means that the time taken to get up and running is much quicker, resulting in the improved turnaround times we have seen.”
In conclusion, senior biomedical scientist Dave Edmondson says: “We have real confidence in the intelliPATH and in Menarini. It is nice to have a system that allows you to walk away without any worries, or the need to check things constantly. The results are excellent and Menarini has provided us with excellent service and support.”