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Filter-Based Plate Readers

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Filter-Based Plate Readers Strapline MONOCHROMAT OR FILTER- BASE D PLATE READERS horses f or c ou rses, or a win ni ng com binat io n? By Dr John Comley This article is based around feedback obtained in HTStec’s recent Multi-Mode Microplate Reader Trends 2007 Report. It highlights findings on what influences reader choice, current use of different plate reader detection modalities, main target types/classes investigated using plate readers, sources of assay reagents, and where different reader types (filter-based vs monochromator) are preferred. The survey found that sensitivity and flexibility in a multi-mode format were the key decision factors in the purchase of a new reader, however both these requirements are difficult to meet in the same machine. Filter-based readers provide maximum sensitivity, also best read speed and can accommodate particular read modes. Monochromator-based readers provide greatest flexibility, wavelength scanning capability and reduced running costs. Vendor updates reviewing their latest multi-mode plate reader developments are discussed together with comments on how their products attempt to address the competing challenges of sensitivity and flexibility. Of particular interest is the emergence of the first hybrid plate reader, ie one that has both monochromator and filter-based detection modules installed within the same instrument, and readers that make use of elements of both components. The report suggests that in the future, a greater proportion of labs will make use of both reader types, lending support to the suggestion that a significant market opportunity now exists for a hybrid plate reader. 2 Drug Discovery World Fall 2007 Strapline icroplate readers, particularly those 2007 market value ($ million) 142 which are able to read multiple detec - Mtion modes (multi-mode) are a very Growth rate (CAGR) 6% common sight in today’s pharma, biotech and aca - demic research laboratories and were the subject of 2007 volume (Units sales/year) 1,700 detailed investigation in HTStec’s recent Multi- Mode Microplate Reader Trends survey and mar - 2007 median reader price range $65,000-$110,000 ket report (published May 2007). MARKET SEGMENTATION: Multi-mode microplate reader market Large pharma 23% In this report the global market for multi-mode microplate readers was estimated to be around Academic research 25% $140 million and growing at 6% annually. With a median price range of $65,000-$110,000, this is Small-medium pharma and all biotech 52% equivalent to sales in excess of 1,700 units a year. This market was segmented by sales value to Table 1: Multi-mode microplate readers – global market estimate approximately ¼ Large Pharma, ¼ Academic Research and ½ Small-Medium Pharma and All Biotech, with sales in the Academic Research seg - ment predominantly in the price range $35,000- $65,000 ( Table 1 ). Monochromator vs filter-based reader – sales and current use Figure 1: Current breakdown of multi-mode plate Multi-mode microplate readers fall into two main categories: 1) filter-based, which use filters to select reader sales the wavelength – two filters are required per appli - Other cation, one for the excitation wavelength and the 11% other for emission wavelength; and 2) monochro - mator-based, which use multiple (sometimes up to Filter-based four per reader) diffraction gratings to create the 50% desired excitation and emission wavelengths, with the wavelengths selected (tunable) through soft - ware. The current breakdown of multi-mode plate Monochromator reader sales was estimated in the report to be 50% 39% filter-based, 39% monochromator-based and 11% other (which includes multi-mode imagers and © HTStec 2007 some speciality single mode readers) ( Figure 1 ). The current lab use of different plate reader types, however, presents a different picture ( Figure 2 ) and shows that the majority of labs still use only filter- Figure 2: Current lab use of different plate based readers (39%) or mainly filter-based readers (29%) today. In comparison the current use of reader types only monochromator-based readers (12%) or Monochromator only mainly monochromator-based readers (7%) is rel - 12% atively small today. Only 13% of survey respon - Mainly Filter-based only 39% dents claimed equal (50:50) use of filter-based and monochromator 7% monochromator-based readers today. 50:50 filter: Use of different detection modalities monochromator The current use of different microplate reader 13% detection modalities is shown in Figure 3 . This Mainly filter-based shows that absorbance (23% share) and fluores - 29% cence intensity (22% share) were the main detec - © HTStec 2007 tion modalities used when the market as a whole Drug Discovery World Fall 2007 3 Strapline was surveyed (ie including the academic segment). Figure 3: Current use of different microplate reader Glow luminescence (13% share) and time-resolved detection modalities FRET (11% share) were then the next most com - monly used detection modalities. A variety of other Absorbance 23% Fluorescence Intensity (FI) 22% readouts (mainly fluorescent), each with a small Glow Luminescence 13% percentage share, make up the remainder of use Time Resolved FRET (TR-FRET or HTRF) 11% Fluorescence Polarisation (FP) 7% today. These other minority readouts also include Radiometric 5% AlphaScreen which is only available on some filter- Time-Resolved Fluorescence (TRF) 4% Fluorescence Resonance Energy Transfer (FRET) 4% based readers; radiometric, which is only rarely AlphaScreen 3% supported on multi-mode readers (eg Hidex Flash/Fast Kinetic Fluorescence (requires injection) 3% Flash Luminescence (requires injection) 1% Chameleon V); and label free which is only avail - Fluorescence Lifetime (FLT) 1% able as a single mode reader (eg Corning Epic). Label Free 1% Plate array based detection 1% What Figure 3 demonstrates is that users make use Flash/Fast Kinetic Absorbance (requires injection) 1% of a wide diversity of read modes and would prefer BRET (ie BRET1 & BRET 2) 0% to see them offered in one instrument today. Those 0.0% 5.0% 10.0% 15.0% 20.0% 25.0% % Use today in 2007 readouts whose use are expected to make the biggest growth over the coming years include time- resolved FRET, flash/fast kinetic fluorescence Figure 4: Main target types and classes investigated (requiring injection) and label free. on plate readers Target classes investigated using microplate readers Enzymes – kinases 21% Receptors – GPCRs 13% The main target types and classes investigated Protein-protein interactions 10% using plate readers alone are presented in Figure 4 . Enzymes – other 9% N/A – Do not investigate targets 9% This shows that enzymes (kinases) (21%) were the Second messengers – cAMP 7% most popular target followed by receptors Enzymes – proteases 5% Enzymes – phosphatases 4% (GPCRs) (13%), protein-protein interactions Second messengers – Ca2+ 4% (10%), enzymes (other) (9%), and then second Receptors – cytokines 4% Protein-DNA interactions 2% messengers (cAMP) (7%). A further 9% of survey Transporters 2% Ion channel – ligand gated 2% respondents indicated they did not investigate any Enzymes – polymerases 1% targets using their plate readers. Overall most plate Protein-peptide interactions 1% Ion channels – voltage gated 1% readers appear to be used to study multiple target Second messengers – IP1/IP3 1% types or classes and as such frequently need to Receptors – nuclear hormone 1% Enzymes – transferases 1% access many different readout technologies. This is Second messengers – GTP 1% further reflected in Figure 5 which shows the pre - 0% 5% 10% 15% 20% 25% ferred source of reagents used in plate reader % Investigating as main target class assays. The relative high use of home brew © HTStec 2007 reagents suggests that many end users, particularly in the academic research segment, still design and Figure 5: Preferred source of reagents in develop their own assays. In comparison, the use of commercial reagent sources appears to show no plate reader assays strongly favoured vendor. Overall, these observa - N/A – Use home brew reagents 24% tions support the view that multiple assay tech - Other reagent supplier 12% nologies are used and there is a need for maximum CisBio 11% flexibility in multi-mode plate readers to support Invitrogen 11% Molecular Devices 11% this diversity of technologies. PerkinElmer (Euroscreen) 10% Promega 7% Why microplate readers are chosen? GE Healthcare 4% The technical factors that most influence the choice Cell Signaling Te chnology 3% Millipore 3% of a microplate reader are ranked in Figure 6 . This DiscoveRx 2% shows that sensitivity and flexibility in a multi- ThermoFisher (Pierce) 2% mode format are the key decision factors, and these Caliper LS (NovaScreen) 0% 0% 5% 10% 15% 20% 25% 30% were ranked above instrument cost. However, % Using as preferred source these requirements tend to be difficult to meet in © HTStec 2007 the same machine. Figure 6 also highlights the fact 4 Drug Discovery World Fall 2007 Strapline that add-ons such as a dispenser capability are nice to have options, but are relatively unimportant in Figure 6: Technical factors influencing the choice purchasing decisions. Reader manufacturers lack - of microplate reader ing a reagent offering are also concerned about the Sensitivity (limits of detection) 4.40 effect this may have on their potential sales. Flexibility (variety of assays types) 4.33 However, data presented in Figure 7 shows that Multimode reader (multiple read-modes offered) 4.29 overall it has little influence on the final decision to Value (cost) 3.90 Ease of use 3.82 purchase a new plate reader. Speed (fast read times per plate) 3.77 Ability to run future assays 3.77 Monochromator vs filter-based Modularity (ability add detection modes in the future) 3.31 Filter-based availability 3.23 reader preference Stacker capability 3.17 Figure 8 attempts to explain under what circum - Temperature control 2.96 stances a particular reader type (ie filter-based ver - Monochromator based availability 2.83 Shaking 2.74 sus monochromator) is currently most preferred, as Dispenser capability 2.48 perceived by end users.
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