Sequencing technology has intensified the mapping of human genome and brings humanity closer to
understand the molecular mechanisms of life. Years after the introduction of the first gene sequencer, gene sequencing technology has played a vital role in enhancing modern molecular research

The completion of the Human Genome Project (HGP) marked the end of ‘the first half of genetics’. Armed with a reference sequence of 3.2 billion bases of DNA, the scientific community has at its fingertips a priceless resource that could change the face of biomedical research and medicine.

The free availability of reference sequence raised a host of crucial questions like, what genetic variations are associated with evolution of mankind? Which genes are associated with complex genetic diseases? What does genetic variation tells us about ethnic susceptibility to drugs and diseases? What is the function of the vast stretches of non-coding or junk DNA? The scientific community across the world quickly realized the need for a paradigm shift in DNA sequencing technology to address these and many other questions. After all, by most estimates, the HGP consumed 13 years and $2.7 billion to map the human genome. It was impossible to reproduce these efforts every time scientists make an attempt to solve the most complex puzzles in medical science; unless it can be done much faster at really affordable cost without compromising the accuracy.

A mighty technology
The DNA  and protein sequencing technology has revolutionized genomics by allowing rapid automated sequencing of genes.  Development in DNA sequencing technology has provided  unprecedented insight into the entire collection of genome transcribed sequences. Without this technology, the progress that led to the biotechnological production of enzymes, active agents as well as antigens for vaccines would not have been possible. The introduction of gene sequencer has extensively accelerated the pace of biological research and various discoveries. The quick pace of sequencing is achieved by employing sophisticated modern DNA sequencing technology.  The technology has helped scientists generate complete DNA sequences of many plants, animals and microbial genomes across the globe. The success of the human genome project has led to rapid advances in the life science industry.

DNA sequencing technology has shown promising results in treating pathogenic infections such as HIV and hepatitis virus, as well as other ailments; and is also gaining prominence as it offers a ray of hope for those afflicted with genetic disorders. The extensive data from protein and DNA sequencing experiments have provided scientists with a wealth of information that forms the basis for the investigation of cellular processes. Towards the end of their research and development efforts, pharmaceutical companies seek to deliver safe and efficient
molecules.

Technological development
The general methodology has changed relatively little since the introduction of the first protein sequencer. However the use of automated equipment to perform multiple cycles has greatly improved the efficacy of  sequencing. Automated protein sequencing is considered as a major development in the field of  biotechnology. Optimization has allowed the determination of extended sequences of very low abundance proteins. Much effort in recent years has been devoted to improve the sensitivity of protein sequencers by using new, highly sensitive methods for identifying amino acids. A large number of upgraded, automated and sensitive DNA sequencing machines continue to enter the market at a much cheaper cost.

Over the last decade significant improvements have been made to DNA sequencing technology. Some of these landmark improvements include shifting from radioactively labeled nucleotides to more safe fluorescent dye labels as well as the move from slab gels to capillary electrophoresis (CE) based analysis.

Even entry-level DNA sequencing platforms currently available in the market completely automate all the sequencing tasks that were time-consuming and laborious. The processes like filling the capillary with gel, sample injection, electrophoresis, real-time detection of labeled DNA strands and converting raw data into analyzed data are now automated. This allows researchers to focus on discovery instead of spending time on the technique itself and downstream data analysis. Further advancements include increased throughput, widely accepted standards for internationally acceptable data and significant reduction in ‘per sample’ cost. These advancements have resulted in the development of increasingly versatile applications, genotyping, SSCP and methylation studies to name a few. The development of more efficient and easy-to-use consumables and efficient sample preparation kits are now a focus area for all the major players in the market.

Since its discovery, PCR has established itself as the most reliable technique that is accelerating biotech research and gaining prominence in almost all areas of life sciences

The discovery of polymerase chain reaction or PCR has forever changed the molecular biology world. It is an indispensable research technique capable of producing billions of copies of a DNA fragment from just few copies, in less than two hours and is used for a variety of medical and biological applications from basic gene sequencing, diagnosis of hereditary diseases, the identification of genetic fingerprints, the detection and diagnosis of infectious diseases to the creation of transgenic organisms.

Following its invention 26 years ago, PCR has been adapted extensively for numerous molecular biology applications. Gene expression analysis by reverse-transcription quantitative PCR (RT-qPCR) has been a key enabling technology of the post-genome era. PCR is also the lone technique that helped the synthetic oligonucleotide business become a thriving industry today.

The invention
Kary Mullis, who earned a PhD in biochemistry from University of California, Berkeley in 1973, conceived PCR as a means to amplify a specific locus of interest on the human genome in 1983 .

After conceptualizing PCR, Kary labored for a number of months to work out experimental conditions. Since thermostable polymerases were not yet available, it was necessary to add Klenow after each thermal cycle, adding to the tedium of development. There were many failures and many reasons why PCR should not work.  Ignoring the doubts of many, the scientist was able to perform his first successful experiment on December 16, 1983. A patent for PCR was awarded to Cetus Corporation, where Mullis worked then. The Taq polymerase enzyme was also covered by patents. Perkin-Elmer partnered with Cetus to commercially introduce a thermal cycler in the market. This platform was based on compressor driven refrigeration technology.  Few years later, pharmaceutical major Hoffman La Roche purchased the rights to the patents in 1992 and currently holds those that are still protected.

Evolution of  technology
As PCR introduced capabilities to identify, manipulate, and amplify DNA, research possibilities flourished. The detection of genetic mutations, the ability to detect the presence of previously unknown genetic material, as well as the ability to analyze degraded DNA, all became common practice. For example, diseases such as muscular dystrophy and HIV could be detected and diagnosed with the use of PCR.

As scientists grew more familiar with the technique of PCR, they began to expand on the utility of the method. And there have been a lot of improvements in the technology, experimental design, and data analysis.

In the late 1980s, PCR was used to measure the quantity of DNA present in a reaction, generating the term “quantitative PCR” or more simply, q-PCR. This technique further improved PCR by the isolation of Taq Polymerase in the early 1990s. qPCR and, more specifically, real-time qPCR has become a routine and robust approach for measuring the expression of genes of interest, validating microarray experiments and monitoring biomarkers. The use of real-time qPCR has nearly supplanted other approaches like Northern blotting and RNase protection assays.

PCR technology and method has today reached a mature stage of development and implementation.  This technique further improved PCR by the isolation of Taq Polymerase in the early 1990s. The heat stable polymerase could remain active through many cycles of heat required for amplification and created the demand for faster cycling. Russell Higuchi and associates developed a system to monitor the amplification of DNA simultaneously to the reaction. The system involved ethidium bromide, a thermal cycler to irradiate samples with UV light, and a camera to record fluorescence.

In the early 1990s, fluorogenic dual labeled probes were developed as a means to practice q-PCR. In conjunction with fluorescent probes, PCR had further evolved into a sensitive quantification tool useful for the detection of any desired genetic element. As a result, the ability to measure gene expression and practice genotyping quickly became trivial and widespread throughout the biotechnology industry. Now, with the recent development of new dyes and quenchers such as the series of Black Hole Quencher, CAL Fluor and Quasar dyes, the possibilities for PCR are seemingly endless.

Market
The real-time PCR market in itself has come a long way, since the introduction of the first commercial real-time PCR platform by Applied Biosystems. The focus has gradually changed from hardware to complete end-to-end workflow-based solution. Gradually, other players also started penetrating the market by introducing more user-friendly and flexible products.

There are over 15 companies operating in this space globally. Applied Biosystems has the highest share of the market with Bio-Rad, Stratagene, Eppendorf and Roche being the other major stake holders in the basic research market. The use of molecular techniques in all segments of the clinical, diagnostic and testing markets has been growing by leaps and bounds over the last few years. This is the primary driver for the growth in the PCR market at present.

Technology upgradations
During the developmental stage of this product line, real-time PCR was perceived merely as an optical upgrade of conventional PCR. But as more sophisticated applications were enabled by the real-time PCR technique, instrument manufacturers also realized that the technique offered great potential and the need for rapid improvements in their platforms. As a result, hardware features including the light source, the detection systems etc. were rapidly upgraded to keep pace with the application development without diluting the cost to performance ratio. Innovative approaches such as virtual filters were also implemented to deconvolute complex fluorescent dye spectra thus achieving better signal to noise ratios.

Even though, there are various real-time PCR chemistries available, two chemistries are widely used by the scientists all over. These are FRET-based TaqMan technology and SYBR Green chemistry which is dependent on SYBR Green’s ability to bind any double-stranded nucleic acid molecule. TaqMan chemistry is extremely accurate and highly specific and considered to be the gold standard for real-time PCR experiments. However, with proper optimization SYBR green based methods can also work well.

The most recent and novel innovation in this area has been the adoption of the microarray format for running real-time PCR experiments. This format allows researchers to interrogate the expression level of hundreds of genes simultaneously. With increasing number of scientists wanting a much closer view of gene expression at the pathway and individual biological process level, which typically involves a few hundred genes, these real-time PCR based low density arrays have already become a very popular tool. The future of PCR remains bright as the technology becomes more rapid, cost-effective, easier to use, and capable of higher throughput.

LC-MS has emerged as the most preferred technique for routine bioanalysis and there is no doubt that this technology will continue towards more automation and wider range of applications in the coming years.

Liquid chromatography-mass spectrometry (LC-MS), the analytical chemistry technique that combines the physical separation capabilities of liquid chromatography with the mass analysis capabilities of mass spectrometry has emerged as a powerful technique used for many applications with very high sensitivity and specificity.

LC-MS are used in innumerable analytical fields, including pesticide-residue determination. There is no doubt that LC-MS is currently competing with gas chromatography (GC)-MS for the status of reference analytical technique to determine pesticide residues and that its ever-increasing application is bound to the evolution of modern instruments and their growing performance qualities. Strategies in the drug discovery and drug development processes are undergoing radical change. For example, the contribution of pharmacokinetics to both processes is increasing. Furthermore, toxicokinetics is established as an essential part of toxicity testing. With this emphasis in the use of pharmacokinetics/toxicokinetics and the greater potencies of newer drugs, a sensitive and specific bioanalytical technique is essential, LC-MS and liquid chromatography-tandem mass spectrometry (LC-MS/MS) have emerged to fulfill that need. LC-MS/MS can be considered to be the major bioanalytical development of this decade.

LC-MS has become very common in pharmacokinetic studies of pharmaceuticals.  The last few years have witnessed its extensive use in the study of proteomics. It is frequently being used in drug development at many different stages including peptide mapping, glycoprotein mapping, natural products dereplication, bioaffinity screening, in vivo drug screening, metabolic stability screening, metabolite identification, impurity identification, degradant identification, quantitative bioanalysis, and quality control.

Technology upgradation
The last few years have seen a significant movement toward using single nanoscale LC-MS   methodologies that are capable of qualitative profiling and quantitative analysis of complex mixtures of proteins across wide concentration ranges and the main focus is in things at the lowest concentrations. Scientists are hence relying on analytical instruments for structural information on individual low-abundance proteins and simultaneously get a true fingerprint of all of the proteins in a given cell. It has become possible for LC-MS techniques to uncover 1,500 proteins in a single analysis.

The major development in MS has been the use of tandem MS-MS where a researcher can program the detector to select certain ions to fragment. The process is essentially a selection technique, but is in fact more complex.  As long as there are no interferences or ion suppression, the LC separation is quite fast. It is common now to have analysis time of one minute or less by MS-MS detection, compared to over 10 minutes with UV detection.

Some of the key improvements in liquid chromatography and mass spectrometry come from better software, such as Waters Corporation’s Micromass ProteinLynx Global Server 2.0, which integrates key mass informatics tools for protein identification and characterization. Such a combination of liquid chromatography, mass spectrometry, and powerful software have proved to be useful in disease target identification and drug discovery.

Future technologies
Although the use of LC-MS for bioanalysis began only a decade ago, growth in the development and applications of this technology has been phenomenal. The performance to cost ratio of the necessary equipment is being improved continuously. The advent of the ion-trap as a tandem MS quantitative detector for routine bioanalysis probably represents a notable breakthrough in terms of performance-cost ratio, despite some compromise in performance such as sensitivity and precision, when compared to triple-quadrupole instruments. Today, instrument manufacturers are more focused on their product range and the equipment is increasingly dedicated to specific applications. Also, with the advances in pumping technology, electronics and software control, instrument manufacturers are able to design machines that are smaller, simpler to use and with a much better performance to cost ratio.

A new generation of LC-MS interfaces that are more amenable to the use of non-volatile buffers and ion pairing reagents are being developed. The technology for automated sample preparation continues to improve. As the range of available packing materials for solid-phase extraction increases, more bioanalytical methods will be based on this extraction approach. In the near future, automated SPE systems or on-line precolumn switching capabilities are expected to become an integral part of a bioanalytical LC-MS/MS system. Other automated SPE methodology will be based on immunoaffinity columns and other molecular recognition approaches. Development of the technology will therefore continue towards more automation and to include an even wider range of applications.

The LIMS technology has given a new insight to scientists for conducting experiments more efficiently and providing greater lab productivity and functionality along with automated reporting capabilities

Laboratory Information Management System (LIMS) is a computer software that is used in the laboratory for the management of samples, laboratory users, instruments, standards and other laboratory functions such as invoicing, plate management, work flow automation etc. LIMS are therefore the information management system designed specifically for the analytical laboratory. One of the most important aspects of LIMS is the ability to prepare and retrieve data and turn it into information quickly and easily. The LIMS has thus been able to eliminate the time consuming tasks of manual report preparation that often preclude access to the desired information.

A full-featured LIMS manages various lab data that ranges from sample log-in to reporting the results. LIMS has streamlined the data flow within organizations and has centralized the information in one primary database. The LIMS solution unifies vast and disparate volumes of biological and chemical data along with their related applications and tools, into a single, browser-based scientific interface. Built mostly on an extensible life science-based data model, the LIMS platforms function on the basis of the context of data being integrated, the relationships between associated data, and allows scientists to use the platform to query, view and analyze research data without reformatting the data-gathering methodology or changing multiple product interfaces.

Today, LIMS has become a comprehensive enterprise solution that enables research organizations to focus their time and effort on production of scientific information. Scientists are now able to leverage and utilize individual work flow within the overall process flow to meet their unique requirements. LIMS has also created a powerful environment in which scientists can interpret their results. Sample data are quickly combined with data integrated from public or proprietary databases by creating a web of information that provides a context for analysis. In addition, integration with the best technologies, platforms and web-based search engines has enabled scientists to quickly transform data into meaningful information.

History
Originally, LIMS were developed in-house by organizations wishing to streamline their data acquisition and reporting processes. In-house LIMS, which are still being developed, take considerable time and resources for its implementation. The need for a more immediate solution helped to drive LIMS to the next stage in the 1970s. During this time, custom-built systems became available. These early custom systems were one-off solutions designed by independent systems development companies to run in specific laboratories. Parallel to these custom-built LIMS implementations were the initial efforts to create commercial LIMS products. These extensive research efforts resulted in the first commercial solutions that were formally introduced in the early 1980s. Such commercial LIMS were proprietary systems, often developed by analytical instrument manufacturers to run on the chromatographs that the instrument  manufacturer produced. These commercial systems, while typically developed for a particular industry such as the pharmaceutical industry, still required considerable customization to meet a specific laboratory’s needs. In particular, laboratories often required very specific format and reporting requirements. However, such demands for customization increased the cost of the commercial LIMS and extended the implementation time. Parallel to the rise in commercial LIMS was the increase in processing speed; the increase in third-party software capabilities; and the reduction in PC, workstation and minicomputer costs. These advantages were transferred to the laboratory and LIMS environment that resulted in a migration from proprietary commercial systems toward an open systems approach that emphasizes user-configurability rather than customization, which took place in
the 1990s.  

Technological development
Today’s commercial LIMS offer a high degree of flexibility and functionality. Many popular commercial LIMS take advantage of open systems architectures and platforms to offer client/server capabilities and enterprise-wide access to lab information. Web-based LIMS, or a web-based front-end to the LIMS, are also offered by many vendors. Extensible Markup Language (XML) is being incorporated into LIMS because it can enhance the information in documents, simplify web automation, and integrate applications within or between organizations. XML not only offers a more streamlined way to transmit data to web applications, but it can also be validated. XML is considered as the next generation LIMS. Informatics is redefining this field. The rise of informatics, coupled with the increasing speed and complexity of the analytical instruments, is driving more sophisticated data manipulation and warehousing tools that work together with LIMS to manage and report laboratory data with ever greater accuracy and efficiency.

Early LIMS were custom-designed and built as point solutions to meet the needs of specific laboratories. This type of in-house systems need significant effort to develop and while they may produce a very close fit to the initial requirements, they can prove difficult to change or modify in response to changing laboratory and business practices. Commercial LIMS products were initially developed in the early 1980s. At first these systems often required significant customization to meet a specific laboratory’s needs as far as work flow and reporting were concerned. This increased the cost of the LIMS setup and extended the implementation time required.

Open LIMS systems
As computing technology changed, LIMS technology developed in parallel and a move towards more open and configurable COTS (Commercial off the Shelf) systems took place. This provided laboratories with greater flexibility to meet their needs within a standard commercial package.

Today’s LIMS solutions offer even greater flexibility and functionality. Many popular commercial LIMS packages utilize open system architecture to offer client capabilities and enterprise-level access to lab data within a client server environment. Some vendors offer the same capability within a truly web-based LIMS using technologies such as Microsoft’s .Net platform. XML enhances the data in documents, maintains data longevity by storing it in an application neutral format, simplifies automation, and integrates data and information within organizations. This, together with the adoption of technologies such as web services, allows for enhanced and simplified integration with other systems within the laboratory or organization. Combined with a fully integrated Scientific Data Management Systems (SDMS), LIMS now has the potential to bring all laboratory data together in a single unified repository.

Next level of development
The impact of informatic technology has always been a bonus for the LIMS industry. Internet, HTML, XML and  hardware improvements have made data management tasks much easier to perform. Portable or wireless devices are part of these trends. Experts predict that wireless technology is on the ascendant at the same time the web-deliverable LIMS are finally gaining market acceptance. The drive to provide even  more functional and productive technology results in easy to use and less expensive LIMS solutions, laboratories worldwide will be able to acquire, manage and report their data in more interesting ways.

DNA microarray has emerged as a prime technology for the performance of gene expression analyses. Combined with bioinformatics and other advanced technology, it offers numerous applications with absolute accuracy.

Availability of whole genomic sequences of many organisms have created the need for high throughput analysis of gene expression patterns and the DNA chip and microarray technology have revolutionized functional and genomic analysis at this level. This technology uses a single chip to  monitor the whole genome so that researchers can have a better picture of the interactions among thousands of genes simultaneously. In the past several years, this new technology has attracted  several biologists as it generates large amount of data in little time and facilitates the quantification of thousands of genes from many samples

DNA microarrays are used to examine the gene expression changes in cancer patients. Tumor profiling, using DNA microarrays, allows the analysis of the development and the progression of complex diseases. The technology allows scientists to examine targets for drug discovery, potential diagnostic and prognostic biomarkers for many complex diseases; detect viruses and other pathogens from blood samples and thus evolved as a pathogen detection method.

DNA microarrays are now used to identify inheritable markers, and therefore used as a genotyping tool. SNP chips based on DNA microarray technology allow the high throughput profiling of single nucleotide polymorphisms using a chip or array approach.  This has allowed polymorphisms to be more quickly assayed and also their relevance to disease to be easily determined. Today, the technology has emerged as an indispensable research tool for gene expression profiling and mutation analysis.

The beginning
Microarray technology has evolved from southern blotting, where fragmented DNA is attached to a substrate and then probed with a known gene or fragment. The use of a collection of distinct DNAs in arrays for expression profiling was first described in 1987, and the arrayed DNAs were used to identify genes whose expression is modulated by interferon. These early gene arrays were made by spotting cDNAs onto filter paper with a pin-spotting device. The use of miniaturized microarrays for gene expression profiling was first reported in 1995, and a complete eukaryotic genome (Saccharomyces cerevisiae) on a microarray was published in 1997. Affymetrix further developed DNA microarrays which were based on high-density 25-mer oligos from human cDNA sequences. Microarrays were originally designed to measure gene expression levels of a few genes. 

Terminologies that have been used in the literature to describe this technology include biochip, DNA chip, DNA microarray, and gene array.

Today, more than 1,000 microarray core facilities are available and over 100 service companies are offering microarray processing services worldwide.

Application advances
As more information accumulates, scientists are able to use microarrays to ask increasingly complex questions to perform more intricate experiments. With new advances, researchers are able to better understand the functions of new genes based on similarities in expression patterns with those of known genes. Ultimately, these studies promise to expand the size of existing gene families, reveal new patterns of coordinated gene expression across gene families, and uncover entirely new categories of genes.

Furthermore, because the product of any one gene usually interacts with those of many others, our understanding of how these genes coordinate will become clearer through such analyses, and precise knowledge of these inter-relationships will emerge. The use of microarrays may also speed up the identification of genes involved in the development of various diseases as it enables scientists to examine a much larger number of genes. This technology will also help in the examination of integration of gene expression and function at the cellular level, reveals how multiple gene products work together to produce physical and chemical responses for both static and changing cellular needs.

Developing new protein arrays and constructing miniaturized flow-through systems, which can potentially take this technology from the research bench into industrial, clinical and other routine applications, exemplify the intense developments that are now ongoing in this field. Recent growth in the field of protein microarray shows the potential applications of enzyme–substrate, DNA–protein and different types of protein–protein interactions. The technology is now more heavily regulated in terms of the bioinformatics, which has led to the generation of more credible results.

Other researchers in the field expect DNA chips to enable clinicians and in some cases even patients to quickly and inexpensively detect the presence of a whole array of genetic diseases and conditions, including AIDS, Alzheimer’s disease, cystic fibrosis, and some forms of cancer. Moreover, the technology could make it possible to conduct widespread disease screening cost-effectively, and to monitor the therapies more effectively. So far, only a few companies have commercialized DNA-chip products, and the barriers to market entry remain great.

Challenges
Microarray technology is expensive in terms of cost of required equipment, reagents and trained manpower. The technology is rapidly advancing; frequent upgradation of machines and methods becomes a major bottle neck in maintaining a microarray lab.

The major hurdle in the efficient utilization of microarray technology is the lack of trained manpower to analyze the microarray data. Expertise in spread sheet and database operations and analysis packages are essential for efficient statistical analysis of data and to find significant patterns.

As DNA-chip companies prepare to bring their products to market, they have to face major technological, manufacturing  and regulatory challenges. The technology trade-offs involve finding ways to increase the number of arrays on a single chip, as well as increasing the rate of production to meet expected demand.

The main challenge involves achieving these parameters at an acceptable cost. In a tight managed-care marketplace that places a premium on technologies that can either show quick savings or more-efficient results, some analysts say that such unit prices will limit the growth of the DNA-chip market.

Future
Microarray technology offers a ray of hope to personalized drugs and molecular diagnostics as it enables global views of biological processes. With technical development that offers increased sensitivity, microarray technology is expected to become an indispensable tool in the fields of biology, biotechnology, drug discovery, and other application areas. If DNA microarray is used for the development of pharmaceutical products, it can considerably reduce the cost and time for the entire process of drug discovery and development, and can also contribute in developing personal drugs.

Although this marketplace is in its infancy, with considerable challenges remaining to be overcome, the speed with which manufacturers are progressing toward commercialization will soon make DNA chips a viable alternatives to traditional chemical assays. Indeed, manufacturers hope that within a decade they will usher in a new era in diagnosis and treatment for diseases and conditions that have genetic origins.

Since its invention, flow cytometry has enabled scientists to analyze a variety of cell types. Today, the applications of this technology are even broader and powerful

Flow cytometry is a technique for counting, examining and sorting microscopic particles suspended in a stream of fluid. It allows simultaneous multiparametric analysis of the physical and chemical characteristics of single cells flowing through an optical and/or electronic detection apparatus.

Early flow cytometers were generally experimental devices, but recent technological advances have created a considerable market for the instrumentation, as well as the reagents used in analysis, such as fluorescently-labeled antibodies and analysis software.

Modern flow cytometers are capable of analyzing several thousand particles every second, in real time, and can actively separate and isolate particles having specified properties. 

The new flow cytometers usually have multiple lasers and fluorescence detectors. Increasing the number of lasers and detectors allow multiple antibody labeling and can more precisely identify a target population by their phenotype. Certain instruments can even take digital images of individual cells and allow analysis of fluorescent signal location within or on the surface of cells.

Fluorescence-Activated Cell Sorting (FACS) has emerged as a specialized type of flow cytometry. It provides a method for sorting a heterogeneous mixture of biological cells into two or more containers based upon the specific light scattering and fluorescent characteristics of each cell. It has become a useful scientific instrument as it provides fast, objective and quantitative recording of fluorescent signals from individual cells as well as physical separation of cells of particular interest.

The invention
The first fluorescence-based flow cytometry device (ICP 11) was developed in 1968 by Wolfgang Göhde from the University of Münster, Germany (patent no. DE1815352) and first commercialized in 1968-69 by German developer and manufacturer Partec through Phywe AG in Göttingen. At that time absorption methods were still widely favored by other scientists over fluorescence methods. The original name of the flow cytometry technology was pulse cytophotometry. After 10 years, in 1978, at the conference of the American engineering foundation in Pensacola, Florida, the name was changed to flow cytometry, a term which quickly became popular. Subsequently Bio/Physics Systems introduced flow cytometry instrument named Cytofluorograph in 1971. In 1973 Partec introduced  PAS 8000. The first FACS instrument from Becton Dickinson came in 1974. ICP 22 from Partec/Phywe and Epics from Coulter were introduced in 1975 and 1977-78 respectively.

Widening applications
The use of flow cytometry has increased considerably during the past decade. The technology has enabled the rapid measurement and analysis of multiple characteristics of single cells. Flow cytometric DNA has been found valuable in determining the biological behavior of various tumors and predicting clinical outcomes. The technology has applications in a number of fields, including molecular biology, pathology, immunology, plant biology and marine biology. In the field of molecular biology it is especially useful when used with fluorescence tagged antibodies. It has broad application in medicine especially in transplantation, hematology, tumor immunology and chemotherapy, genetics and sperm sorting for sex preselection. In marine biology, the auto-fluorescent properties of photosynthetic plankton can be exploited by flow cytometry in order to characterize abundance and community structure. In protein engineering, flow cytometry is used in conjunction with yeast display and bacterial display to identify cell surface-displayed protein variants with desired properties.

Mr Ram Sharma, managing director, BD India, said, “We never imagined that flow cytometry will become core for monitoring CD4. This is now the most preferred solution for monitoring HIV/AIDS patients. Almost 90 percent of all CD4 monitoring is done using our flow cytometry. Although the technology already has a lot of clinical applications in monitoring cancer, HIV, cord blood banking, and stem cells, it has not been intensely deployed in the areas of drug discovery, life science research and basic research. Through partnerships with research institutes, we can increase the scope of our products in the life science research applications. Our primary aim lies in enhancing our scope in drug discovery.”

Market overview
Compared to other technologies, there are few players operating in the flow cytometry market space, the important ones being Beckman Coulter, Guava Technologies, Luminex and Dako. Talking about market competition, Mr Sharma said, “Over a period of time, the technology becomes less competitive. The real difference is the quality of people who are training and educating the customers and the support that you provide to your customers. The manufacturers have to keep a constant check on the efficiency of the systems and probability of new applications for
investment.”

The flow cytometry market is composed of instruments, reagents, devices, and services used across the research and clinical life sciences areas that span the academic and biomedical, biotechnology, and pharmaceutical business market sectors. According to a market research report, the global flow cytometry market in 2008 stood at $1.5 billion and is estimated to grow to $3.7 billion by 2015 with estimated growth rates of existing product. About 68 percent of the 2008 product revenue has been from instruments, and reagents make up 32 percent of the revenue. Cell-based flow cytometry is estimated at $1.3 billion with a CAGR of 10-15 percent in the coming year. Market leaders Becton Dickinson and Beckman Coulter account for about 70 percent of the research and clinical areas of the cell-based flow cytometry market. Bead-based flow cytometry is estimated at $215 million with a CAGR of 25-30 percent while Luminex and partners comprise approximately 90 percent of the bead-based flow cytometry market.

Anticipated developments
Researchers expects more fluorescent dyes to become available, with more color options, and also further technical improvements. These include dyes with much higher stoke shifts and alternatives for tandem dyes, which often suffer from inherent problems in multi-laser applications. Upgradation of some existing dyes are in the pipeline, which will offer increased brightness for conjugated antibodies. It is also predicted that new dyes will be offered together with new laser options from flow cytometer manufacturers, with an emphasis on accessibility. New solid state lasers in digital instruments are providing options in bench top machines that were previously restricted to top-end instruments.