The Companies Driving it To Market: Slide Free Microscopy Part 3
The Companies Bringing Slide-Free Microscopy
After exploring why slide-free microscopy holds so much disruptive potential in Part 1 and the possible applications for these technologies in Part 2, it’s now time for the one everyone’s waiting for: the people and the companies actually making it happen. In this third installment, we’ll peek behind the curtain at four startups, each bringing their own unique approach to slide-free microscopy. There’s much more we could say about each, but I’ll leave that to future newsletters and your own exploration. Loads of colorful photos and videos are included below, so read on!
Company #1: Alpenglow Biosciences
Alpenglow was founded by pathologist and CEO Nicholas Reder MD MPH. Interestingly, Nick was one of the early inspirations for my excitement about slide free microscopy. Back in 2017, I ran across a story about Nick and his team when he was a resident at the University of Washington being recognized by NIH Director Francis Collins for his lab’s groundbreaking technology. Nick’s work at the University of Washington led to the development of a patented open-top light-sheet microscope technology that formed the basis of the company, Alpenglow Biosciences. (Side note: I think Alpenglow has one of the coolest names for an imaging company! Go look up the definition)
How it Works
Alpenglow uses open-top light-sheet microscope technology to image tissue, combining imaging with an AI-enabled cloud-based computing and analysis system to produce 3D spatial analysis of tissue samples at high speed. The system can be used to analyze entire tissue samples or conventional multi-well plates.
To give you a taste of how cool these 3D images are, check this video out:
The workflow
- Fixation: Preferably 4% PFA (although formalin works too).
- Pre-treatment: Depending on tissue type, there may need to be bleaching, delipidation, or decalcification
- Staining: Either small molecule fluorophores or antibody staining. Antibody staining involves antigen retrieval, primary labeling, and secondary labeling.
- Dehydration: Run through graded ethanol ladder
- Clearing: Immersion in a clearing agent such as ethyl cinnamate
- Imaging: Tissue is placed in transparent refractive index-matched specimen holders. Z-stacked optical sections are acquired using the Aurora 3Di hybrid open-top light sheet microscope.
- Image Viewing: Images are processed using the Alpenglow processing pipeline and then ready to view.
What’s unique about Alpenglow’s technology?
- True 3D Imaging
Provides spatial context for pathology, letting one visualize tissue as a volumetric structure rather than just a single plane.
- AI and Machine Learning
More data generated from 3D imaging means more opportunity for machine learning models
- Multiple Immunomarkers
The ability to stain for multiple markers in the same sample helps pinpoint specific cells or structures of interest.
- Just freakin’ cool
The 3D images are stunning. Navigating them feels like we’re entering a new era of imaging, blending pathology with radiology.
Potential Applications
- Enhanced Research
By enabling high-speed, AI-powered 3D spatial analysis of tissues, Alpenglow opens a whole new layer of discovery. In preclinical studies, this approach could provide a detailed view of drug localization, visualization of complex structures like vessels and nerves, and identify subtleties overlooked in 2D approaches.
- Drug Trials
Alpenglow’s system opens up new opportunities for mechanism of action studies for drug trials.
- Rapid Surgical Guidance
Although not a big focus of the company now, the technology may be capable of providing rapid surgical guidance in the future.
Combining 3D Spatial Analysis with Targeted Evaluation
Alpenglow stands out in its ability to generate incredible 3 dimensional images that give us a new look at what tissues look like under the microscope. Complex anatomical features, like nerves weaving through tissue, can be evaluated in three dimensions. The ability to stain for and visualize multiple markers in a tissue allows us to hone in on structures of interest. Machine learning combined with Alpenglow’s new type of three dimensional microscopy data open up exciting areas of research and exploration.
See some more examples of Alpenglow’s images below. Check out one of their informational webinars here if you’d like to go deeper.
Company #2: Applikate Technologies
I remember the first time I saw an image generated by Applikate. I’d been watching these various techniques for a few years. Most of the images I’d seen were cool and promising, but were still far from diagnostically useful. The few high quality imaging techniques took too long or were too expensive to be brought over to clinical diagnostics. Then I saw my first image from Applikate, which just happened to be the image of the glomerulus in the gallery below, and got really excited about the potential for slide-free microscopy coming to clinical diagnostics. Founded by Richard Torres and Michael Levene, Applikate Technologies is a New Haven, Connecticut-based hardware and software company. Applikate brings a unique and patented approach to multiphoton microscopy that offers an automated histology platform with digital imaging in a benchtop device. Michael Levene, PhD (in computation and neural systems), spent a decade on Yale’s faculty (2004–2014) pioneering the fusion of multiphoton microscopy with optical tissue clearing before co-founding Applikate. Richard Torres, MD, is a pathologist and biomedical engineer with deep expertise in medical lab instrumentation. He co-invented Applikate’s novel histology techniques and devices, served on Yale’s faculty as well, and joined forces with Michael to form Applikate.
How It Works
Applikate uses clearing histology with multiphoton microscopy (CHiMP) for tissue processing and image acquisition.
The Workflow
- Grossing
Fresh or formalin fixed tissue is sliced to the standard thickness for tissue cassettes (3mm is ideal). - Processing and Staining
Tissue is dehydrated and stained for 15 minutes-2 hours and then cleared for 5-30 minutes. Total time needed depends on the application and desired quality of image. - Imaging
After processing, the tissue is placed in Applikate’s proprietary tissue holder, placed in the imaging unit, and within minutes a virtual “slide” is generated. The user can pick single-plane scans or go for multiple Z-stack levels if more depth is needed. - Image Viewing
Digital images appear in STACKstreamer, Applikate’s web-based software that lets the user navigate multiple Z-stack levels up to 1.5 mm deep into the sample. Images can also be opened in other commonly used viewing software platforms.
What’s Unique About Applikate’s Technology?
- High Quality images that mimic traditional H&E
The images are designed to mimic traditional H&E slides and are instantly familiar to new users. Resolution and quality are high, with crisp visualization of subcellular structures like nucleoli all the way to bacteria. - Deeper Imaging
With Applikate’s device, you can scan multiple Z-levels into the tissue (up to 1.5 mm deep), replacing the need for deeper serial sections and flats of slides. - 3D Reconstructions
Stacking multiple images for 3D reconstruction opens new doors for analyzing complex tissue architecture that’s difficult to grasp on a single 2D plane. - Fresh Tissue Imaging
Tissue can be imaged fresh, preserving molecular diagnostic opportunities.
Potential Applications
- Histology Lab Replacement
Think of this as a consolidated lab device that squeezes most of the conventional tissue-processing workflow and personnel required into one box. That means faster turnaround times, fewer staff required, and less equipment to maintain. - Point-of-Care Device
Intra-operative or immediate post-operative imaging could be done on fresh surgical tissue, providing real-time feedback in the OR. Imagine not having to wait days for pathology results. All with images that are better than current rapid methods such as frozen sectioning. - Remote Deployments
A small, mobile instrument that can land in satellite labs, resource-limited sites, or emergency-response units. In short, a pop-up histology lab that fits in a box. - Enhanced Research
A device that allows individual research labs to generate pathology images from tissue, without having to rely on external histology labs and their associated costs. Additionally, the potential for 3D reconstruction allowing spatial analysis, combined with machine learning models, opens up exciting opportunities for advancing research.
Balancing Quality with Workflow Simplicity
Where Applikate really shines is in how it marries a simple, straightforward workflow with the kind of high diagnostic image quality pathologists expect, in a fraction of the time required in the traditional process. This combination makes the Applikate device a strong contender for revolutionizing how we generate histopathology images in the lab at scale.
Check out some of these images from Applikate!
Company #3: MUSE Microscopy
A few years after completing the commercial diagnostic workflow conversion to fully digital slide scanning at Antech Diagnostics (read about that conversion here), Dr. Jeff Edwards and I attended the Digital Pathology World Congress in New York City. Much of the conference centered on the typical challenges facing human pathology in making the digital shift, but one speaker in particular caught our attention. Dr. Richard Levenson MD, FCAP introduced a novel, slide-free microscopy technique called MUSE (Microscopy with Ultraviolet Surface Excitation). While the image resolution wasn’t yet on par with traditional H&E, the unique aspect of MUSE was and continues to be its rapid workflow and low-cost imaging device. You can stain fresh tissue in minutes, and proceed directly to imaging, making MUSE a strong candidate for point-of-care use.
How It Works
MUSE utilizes short wavelength UV light to excite fluorescent histology stains, such as DAPI and eosin in the most superficial (5-15 micron thick) layer in tissue, capturing those signals and converting them into digital images.
The workflow
- Grossing: Tissue, fresh or formalin fixed, is grossed to standard cassette thickness.
- Staining: A quick, non-destructive stain is applied ( 1-2 minutes)
- Imaging: Tissue is loaded into a proprietary SmartPath Dx cassette and loaded into the MUSE device. Imaging is rapid, with images viewable in minutes.
What’s unique about MUSE’s technology?
- Rapid Workflow
The tissue prep and staining process is rapid, taking less than 5 minutes to prepare, stain and insert the tissue into the imaging cassette prior to imaging.
- Low Instrument Cost
The instrument price is relatively inexpensive compared to other techniques.
- Surface topography is visible
Surface topography can be captured, since shaped structures on or near the surface of the specimen can be appreciated.
- Preserves Molecular Diagnostics
The non-destructive nature of the tissue prep and staining process preserves molecular diagnostic opportunities.
Potential applications:
- Point-of-care instrument
The inexpensive nature of the instrument along with the rapid workflow makes MUSE a candidate for real-time, intra-aoperative tissue evaluation, or immediate post-operative review in hospital settings.
- Replacing cryosectioning
MUSE could potentially replace cryosectioning workflows, offering simpler prep, minimal training, and less equipment overhead.
- Enhanced Research
AI analysis of surface topography and staining features may reveal novel biology or histopathology.
Muse merges rapid imaging with an affordable device
MUSE’s real strength lies in combining rapid tissue prep with a low-cost imaging device that requires minimal technical expertise. The potential for point-of-care deployment and replacement of cryosectioning applications makes MUSE an exciting technology to watch in the future.
Check out some images from Muse:
Company #4: Histolix
Histolix is a company founded by Richard Levenson,MD and Farzad Fereidouni, PhD in 2019 that takes an imaging approach similar to MUSE, termed “FIBI” (Fluorescence-Imitating Brightfield Microscopy). Like MUSE, the rapid workflow and imaging capabilities enable FIBI to be a candidate for point-of-care histopathology.
How It Works
FIBI employs an epifluorescence microscope with 405-nm excitation. Unlike MUSE, which images signals from fluorescent stains, FIBI uses a combination of standard–absorbing–histology stains such as hematoxylin and eosin. Stained structures are back-illuminated by broad-spectrum tissue autofluorescence generated by the 405-nm excitation and arising from relatively deep inside the specimen. The signals are then captured using an RGB camera, to generate digital images that closely resemble traditional H&E-stained slides.
The Workflow:
- Grossing: Tissue, fresh or formalin fixed, is grossed to standard cassette thickness.
- Staining: A quick, non-destructive stain is applied ( 1-2 minutes)
- Imaging: Tissue is loaded into a proprietary tissue holder and transferred into the benchtop instrument containing the epifluorescence microscope. Imaging is rapid, with images viewable in minutes.
What’s unique about Histolix’s technology?
- Rapid Workflow
The tissue prep and staining process is rapid, taking only 5 minutes of staining before tissue can then be imaged.
- Low Instrument Cost
The instrument price is relatively inexpensive compared to other techniques.
- Preserves Molecular Diagnostics
The non-destructive nature of the tissue prep and staining process preserves molecular diagnostic opportunities.
Potential applications:
- Point-of-care instrument
The inexpensive nature of the instrument along with the rapid workflow makes FIBI a candidate for real-time, intra-operative tissue evaluation, or immediate post-operative review in hospital settings.
- Replacing cryosectioning
FIBI could potentially replace cryosectioning workflows, offering simpler prep, minimal training, and less equipment overhead.
- Enhanced Research
Novel insights into continuous structures on or just below the tissue surface, such as nerves and blood vessels can power new insights into biology or histopathology.
Histolix, another contender for point-of-care slide free microscopy
FIBI is very similar to MUSE in terms of technological approach and potential applications. The ability to combine a low-cost imaging device with a rapid simple workflow opens up exciting possibilities for point of care applications.
Check out these images from Histolix:
Summary
I hope this was a helpful side by side comparison of some of the companies I’m most excited about in the slide-free microscopy arena. Did I miss any that you think should be included? Please message me below. I love feedback and helpful input from my readers.
Disclaimer: I am not currently employed by or affiliated with any company developing slide-free microscopy technology. However, I previously served as a scientific advisor for Smart Health Dx and hold equity in that company. My opinions in this article are my own and are based on my experience in pathology and my independent analysis of the field, and do not necessarily reflect the official views or positions of any companies mentioned. This newsletter is intended for informational purposes only and does not constitute financial, medical, or investment advice.
