MATTHIAS SCHULZEWALLACE LATIMER

Biotech instrumentation manufactur-ers are currently developing productsthat feature increased miniaturiza-

tion and lower costs per use, mainly due toworldwide trends in healthcare. For instru-ments based on laser-excited fluorescence,this has created demand for laser modulesthat emphasize low cost, compact packag-ing and reliability, rather than cutting edgeperformance. This article reviews why andhow these market needs are being met, andalso explores the advantages of lasersources over LEDs for these applications.

Fluorescence-BasedAnalyzers

Many instruments that analyze biochemi-cal reactions and properties utilize fluo-rescent probes which are tagged to spe-cific cells, antigens or sub-cellular compo-

nents (or which are expressed by geneticmodification of the cells themselves). Ex-amples include cytometry, genetic se-quencing, hematology, polymerase chainreaction (PCR), high-throughput drugscreening, and microarray scanners. Theadvantages of fluorescence detection overchemical interrogation are fast, non-con-tact measurements with superb spatialdiscrimination. Plus, these techniqueslend themselves to miniaturization, au-tomation and simultaneous parallel pro-cessing.

Nearly all these instruments use one ormore lasers to excite the fluorescence,where the laser is focused to a small area

or spot size (typical spot sizes are givenin Table A). By using several laser wave-lengths to excite multiple fluorophores,some of these instruments can analyzemultiple cell types in the same data run.

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Laser versus LEDOPTIMIZING LASERS FOR COST-SENSITIVE BIO APPLICATIONS

A new generation of compact, OEM lasers provides the performance and economy needed for

important bioinstrumentation sectors, such as gene sequencers and desktop analytical

instruments intended for point of care.

C O N TA C T

Coherent (Deutschland) GmbH64807 Dieburg, GermanyTel. +49 6071 968-0sales.germany@coherent.dewww.coherent.deLasys: Stand 4-D56

A Typical spot sizes for floureszenze applications

Application Focused Spot Size

Gene Sequencing/Medical Diagnostics 1 to 10 x 1 to 10 mm

Medical Diagnostics 100’s x 100’s µm

Cytometry 30 x 30 µm

Microfluidics andminiaturization lower costs

In many applications, there is shift fromthe research laboratory to the clinical lab-oratory, and, thence, to the point of care.In the West, the goals are personalizedmedicine and lower costs. Personalizedmedicine means genetic profiling of thepatient and/or the specific disease to de-termine what drugs will yield an optimumresult. In the fast growing Chinese mar-ket, the aim is to enable more testingthrough lower costs, bringing the stan-dard of healthcare closer to that of theWest.

In fluorescence-enabled diagnostics,this trend manifests itself in greaterstreamlining and automation. While in re-search and clinical laboratories the usersmay possess expertise in fluorophorechemistry, this is not the case in a doctor’soffice. As a result, the market needs instru-ments configured for simplified and repet-itive use in common tests, rather than themultifaceted flexibility of a typical flow cy-tometer. This trend dovetails with the in-creasing use of microfluidic instruments,often called ›lab on a chip‹, which also en-able much smaller instruments than arepossible with legacy flow technology

Lasers vs. LEDs

High end instruments can easily cost sev-eral hundred thousand Euros, so the use of

one or more lasers to enable high perform-ance is easily justified. But, the develop-ment of miniaturized, lower cost instru-ments is causing manufacturers to askwhether it is cost effective to even use alaser at all. Can some of these applica-tions instead be serviced with a lower-cost LED or super-luminescent LED (SLED)as the fluorescence excitation source?

Many applications don’t need the ex-treme monochromaticity and spatialbrightness of a laser, and a few instru-ments that use very wide-field illumina-tion may be able to economize by usingLEDs. But blood analyzers and lab on achip instruments require a focused spot oflight with a narrow, well-defined and con-sistently reproducible spectral bandwidthand intensity profile. This is actually quitedifficult and highly inefficient to achievewith LEDs, and requires extra optics andcareful LED screening (selection). Thesesteps can push the real cost of implement-ing LEDs above that of using ›entry-level‹lasers.

As one example of the advantages oflasers, note that a 1 W laser can easily befocused to a diffraction-limited spot con-taining over 0.9 W, while the focused spotfrom a 1 W LED might contain only100 µW of useable photons. Part of theproblem is that LEDs are rated by theirpower consumption, not their optical out-put. So, a 1 W-rated LED might only gen-erate a total of 90 mW. Moreover, the LEDemits into a large solid angle, and high

numerical aperture (i.e.,high cost) optics are re-quired to capture just apart of this output. Plus,the LED is not a true pointsource, as is a laser, sothere is an inescapable op-tical trade-off (called›etendue‹) between collec-tion efficiency and finalspot size.

In addition, LEDs out-put a broad spectrum thatusually must be wave-length filtered to avoidcross-talk between differ-ent fluorophore signals, aswell as to minimize back-ground noise due to scat-tered excitation light (Fig-ure 1). All this adds costand complexity to the in-strument, while reducing

the amount of light reaching the interac-tion zone (which can further reduce in-strument signal-to-noise ratio).

Lastly, LEDs are volume manufacturedon a massive scale. The instrument manu-facturer, and even the LED reseller, has nocontrol over the unit-to-unit variations inLED output spectrum and spatial illumina-tion distribution. This presents a seriouschallenge to achieving instrument consis-tency and serviceability.

Optimized Lasers

Of course, the choice of a laser as the lightsource in an analyzer is also critically im-pacted by the cost/performance ratio ofthe available laser products. It is vitallyimportant that the laser offers only thefunctions and performance parametersneeded for optimum instrument opera-tion. Higher laser specifications will in-crease cost without delivering any tangi-ble benefit to the application. To meetthis need, laser manufacturers have intro-duced ›entry level‹ products, such as thenew Coherent BioRay series (Figure 2),that are optimized specifically for thisnew generation of analyzers.

These lasers deliver a few tens of mil-liwatts, and are available at several visi-ble wavelengths which match the opti-mum excitation of common fluorescentprobes and genetically encoded markers(typically 405, 450, 488, 520 and640 nm). These products are based on

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1 Every fluorophore is characterized by an excitation spectrum and a long wavelength emission spectrum. With asingle wavelength laser, it is easy to separate scattered excitation light from the emission spectrum using a long-wave cutoff filter. But with a typical LED, the long wavelength tail of its output overlaps the fluorescence emissionand must be somehow eliminated. Plus the size and shape of this tail can vary significantly between differentbatches of LEDs

Summary

For the fast growing point of care mar-ket, biotech instrument manufacturersneed lasers that meet target perform-ance parameters to achieve the neces-sary performance. But they must alsoavoid the use of lasers that far exceedthese parameters as this unnecessarilydrives up measurement cost. A new gen-eration of lasers has been developed tospecifically match this price/perform-ance sweet spot and become the moreeconomical solution compared to LED al-ternatives.

AUTHORSMATTHIAS SCHULZE is Director of Marketing OEM Com-

ponents & Instrumentation at Coherent Inc.

WALLACE LATIMER is responsible for the Product linie

Machine Vision at Coherent Inc.

n www.laser-photonik.de/LP110256

laser diode technology, as that is thesimplest and lowest cost method of gen-erating CW laser output at these wave-lengths and in this power range.Laser diodes also offer the highestefficiency, lowering the requiredpower budget for the final instru-ment. Since edge emitting laserdiodes emit a highly divergent andasymmetric (elliptical) beam, optics areused in the laser head to produce a colli-mated, elliptical beam. To reduce thecomplexity of downstream beam deliv-ery optics, each head has an ad-justable output lens to enable smoothadjustment of the beam waist location.

These lasers produce a clean Gaussian,TEM00 beam (M2 < 1.5) enabling focusingto a small spot. In comparison lasers withhigher order modes are not well-suited foruse in biotech instrumentation where ad-ditional spatial filtering would be re-quired. Another important feature for in-strument manufacturers is direct analogmodulation up to 500 kHz, in constantpower mode, which supports fast cellcounting and other applications. (Earlierdiode-based lasers only enable modula-tion in constant diode current mode which

can require anextra level of signal normalization.) Twoother advantages are a simple, commonelectronic interface offering both RS 232and GUI control, and a common mechani-cal platform irrespective of output poweror wavelength. This enables simple fieldreplacement or upgrading in the field, of-ten called ›hot-swapping‹.

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2 A new generation of economicalCW visible lasers such as the Coherent

BioRay series has been developed with out-put parameters tailored specifically to meet

current and future trends in biotechinstrumentation