The FACS: A Tricorder for Biologists

Rayleigh, Coulter, Gohde, Swift and Herzenberg—no, not a law firm. These people occupy points on a graph that plots the evolution of a device for medical diagnosis that, for hematologists anyway,  is a bit like a (non-portable) Tricorder. Known as a FACS machine (Fluorescent Activated Cell Sorter), it is a technology so advanced that, as Arthur C. Clarke would say, it is “indistinguishable from magic.”

The Tricorder of course is the hand-held medical device used by the medics in Star Trek. It can diagnose any medical condition with a quick wave. To be honest, a FACS machine stays put, tethered to lasers, tubing, monitors and computers. But give it a suspension of cells to imbibe and it will trickle them through shiny lights and create a plot of the cell types contained therein. From this display the pathologist can, usually, say with certainty that your patient has this or that type of hematologic malignancy, even if it was a mystery to the microscopist, let alone the hematologist at the bedside, who, up to this point, knew little more than the white cells were too numerous, looked odd or the lymph nodes were too big. 

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For decades light microscopy was the only way to enumerate or classify blood cells.  Then the great Max Wintrobe invented the Wintrobe tube. The thin tube was loaded with blood and the cells allowed to settle. The pattern of sorted cells were read for meaning—anemia, inflammation, etc. The problem for modern hematology is more demanding, with many subtypes, often subtle, of white cell diseases. Some are chronic and handled readily with a handful of cheap generic pills. Others are imminently life-threatening and treatment may consist of infusions costing a quarter of a million dollars or more.

Wintrobe Tube

Wintrobe Tube

So if someone were to say, “Here, I have a tool that will tell you exactly which of dozens of different hematologic conditions you’re dealing with. All I need is a suspension of cells and a couple of minutes—be right back," then they’d be talking about a device that borrowed from the brilliance of a nineteenth century nobleman investigating water droplets, and inkjet engineers, animals that glow, and lasers that shine coherently. Before the FACS method was eventually sold by a university to private enterprise, it underwent a long period of incubation. 

We start with the remarkable genius, there’s no other word for him, of John William Strutt, who just happened to be the 3rd Lord of Rayleigh. Lord Rayleigh explained why the sky is blue and how seabirds soar. He won the Noble Prize in 1904 for jointly discovering Argon. Amongst the many natural phenomena bearing his name is Platueau-Rayleigh Instability, in which we learn that a falling stream of fluid will reduce its surface tension by breaking into droplets.

Lord Rayleigh

Lord Rayleigh

Plateau-Rayleigh Instability

Plateau-Rayleigh Instability

Exploiting the flow of droplets to contain individual cells for analysis proved problematic to the biologists working on the prototypes of the FACS. But flinging droplets with great precision had become de rigueur for some of their colleagues at Stanford University—engineers in the process of inventing the ink jet printer. Amongst them was Richard Sweet, author of[ http://www.printhead911.com/inkjet-history/] High frequency recording with electrostatically deflected ink-jets.

When Richard Sweet died, his obit in the San Jose Mercury News (now the journal of record for Silicon Valley) said he had been an avid fisherman and poker player—and the inventor of the inkjet printer. He had five patents to his name, three assigned to Stanford, one to Xerox and one to Becton-Dickinson Company, which bought the FACS technology from Stanford. 

Richard Sweet

Richard Sweet

Leonard Herzenberg

Leonard Herzenberg

The story I heard of how Sweet was recruited to work on the FACS is that he happened at lunch to encounter Leonard Herzenberg, a Stanford geneticist tired of hand–counting cells in his research. He wanted a better way, but truth be told, it went like this: Herzenberg got wind of a possible solution and went to visit Mack Fulwyler at Los Alamos National Laboratory, where Fulwyler had made a working sorter to harvest cell-size particulates from the lungs of rats subjected to radioactive fallout. He got the idea to use electrostatic deflection of droplets from—that’s right—Dr. Sweet. Herzenberg knew just whom to contact back at Stanford.

Their collaboration and $14,000 got them, a few years later, in 1974, “the Whizzer”—the “first” working fluorescent cytometer that could “sort” live fluorescent cells. This created a stir when published in Scientific American. His team recounted the whole story years later in Clinical Chemistry: The History and Future of the Fluorescence Activated Cell Sorter and Flow Cytometry: A View from Stanford.

But innovation didn’t stop there. He and his wife, with whom he collaborated for decades, went to Cambridge on sabbatical. There they learned from Cesar Millstein about monoclonal antibodies. These they adapted to the FACS methodology, thus pioneering a new era of exquisite specificity in biologic diagnostics. 

Herzenberg in the lab

Herzenberg in the lab

Leonard and Lenonore

Leonard and Lenonore

Herzenberg died of a stroke at 81 in 2013. The tribute from Stanford School of Medicine recalls his scientific achievements, including the esteemed Kyoto Prize. They also carefully note that he and his wife and colleague, Leonore, had been active against McCarthy-ism and fostered the role of women and minorities in science. When one of their children was born with Down’s Syndrome they applied FACS technology to develop a means of non-invasive pre-natal diagnosis. When AIDS surfaced amongst friends of his lab, they developed a FACS method for tracking T-Cells in that condition. 

From Scientific American

From Scientific American

Modern Mulitcolor FACS Image from Rebecca Lee

Modern Mulitcolor FACS Image from Rebecca Lee

Herzenberg’s success was, of course, built upon the work of others. Another Stanford researcher, Lou Kamentsky, had built a Rapid Cell Spectrophotometer to cull out cancer cells. He used fluorescence and an Argon—discovered by Lord Rayleigh as we know— laser to do the job. 

Wolfgang Gohde

Wolfgang Gohde

U. Munster–Cells in Motion program

U. Munster–Cells in Motion program

And in Germany, Wolfgang Göhde at Munster University designed a fluorescence based flow cytometer (ICPII) in 1968, commercialized by Partec. Munster University to this day has an extensive program in cell sorting and imaging, called Cells in Motion. He is now at Purdue University, where you can find an extensive history of the German contributions. Volume 10 of the Purdue Cytometry Disc Series is devoted to the history of cytometry.

And before that, Wallace Coulter, in 1949, devised a way of counting red cells flowing in a tube, based on electrostatic changes in the pathway—the “Coulter Principle”. Prior to that Moldavan, in 1934, used a photoelectric apparatus to count red cells as they marched through a capillary tube mounted on a microscope stage.

One can find a Coulter counter in hematology offices and every hospital and commercial lab. His contribution to diagnostic hematology has made of him an icon of biomedical engineering. You can see an interactive timeline of his life on the Wallace H. Coulter Foundation site. To give him his due, let’s just say that his legacy is part of why, in 2017, the Georgia Institute of Technology’s undergraduate biomedical engineering program was ranked #1 in the country, again.

Georgia Institute of Technology

Georgia Institute of Technology

For a short overview of the history of the FACS and its many applications see this article from Microbial and Biochemical Technology. For the real nitty-gritty, check out the Purdue history noted above. 

Labome has a nice survey of how the FACS works, and Clinical Flow Wiki has some canonical images of pathological patterns

Normal Cells

Normal Cells

Acute Myelogenous Leukemia

Acute Myelogenous Leukemia

And, if you must, here’s a nice review of the history of inkjet printing

inkjet-age.jpg

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