Tom Villani Featured in Biocompare Article on 3D Cell Cultures

Check out this recent Biocompare article by Lauren Tanabe, Ph.D. where Visikol Inc's CSO Tom Villani discusses 3D cell culture models and their use in the drug discovery process. 3D cell culture models are being adopted in the drug discovery space at a rapid rate due to their in vivo relevancy. Visikol is leveraging 3D cell culture models and their patented Visikol HISTO technology to offer best-in-class 3D drug discovery assays: 

  • Antibody penetration
  • Apoptosis
  • Angiogenesis
  • Cell-cell junctions and cadherins
  • Cell proliferation
  • Cell migration
  • Cell viability
  • Cholestasis
  • Drug induced liver injury

For more information go to: Services

Visikol HISTO Microtissue
 
 
  • Fibrosis
  • Immune cell invasion
  • Inflammatory Signaling
  • Mitochondrial Health + Toxicity
  • NASH
  • Neuronal Differentiation
  • Neuronal Function
  • ROS
  • Steatosis

 

 

Visikol Presents at Allergan Innovation Forum on 3D Drug Discovery Assays

Recently, Visikol began offering drug discovery assay development services to clients that are focused on 3D in vitro cell culture screening and whole mount tissue analysis. In early September, Tom Villani and Michael Johnson were invited to give a seminar at an Allergan Innovation Forum in Irvine, California about these services and how they fit into the drug discovery process. 

For more information on our services check out Visikol Services or email us at info@visikol.com 

Visikol Gets Second Place at Boston Biotech Week Pitch Competition

This past week Visikol participated in Boston Biotech Week and Dr. Michael Johnson pitched during the biotech startup competition and came in second place overall out of twenty companies.

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The competition brought out companies from around-the-world and was judged by Investors and Business Development Directors from Abbvie, Atlas Venture, Agent Capital, SR One, MRL Venture Fund and Lightstone Ventures.

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Visikol Pitch Competition Boston

Visikol HISTO Assay Services

Our long-term goals as a company are to improve the drug discovery process and the diagnosis of disease. This quarter, we are excited to launch our offering of assay development services for microtissues in drug discovery by leveraging our Visikol HISTO technology. Currently, high content imaging of microtissues introduces an unavoidable bias, since microtissues are too thick and opaque, and thus only the outermost cells can be detected. The outermost cells are more exposed to nutrients, oxygen, drug compounds, etc. with respect to the innermost cells, and thus characterization of microtissues has been fundamentally limited. Using Visikol HISTO, microtissues are rapidly rendered transparent allowing for the characterization of effects of drug candidates on the complete population of cells contained within microtissues. With Visikol HISTO, you can break down the effects of a drug compound according to populations of cells segregated by cell-type, location in the microtissue, etc.

For more information on our drug discovery services and to see if your research question can be answered with our platform, email us at info@visikol.com or check out Visikol Services

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Visikol HISTO Tutorial Videos

We keep in contact with our sample testers and customers to understand how we can improve our user experience so that every customer can incorporate the Visikol HISTO technology into their workflow without friction. Through this process, we identified that many of our customers wanted more detail and background on how exactly our products work and a step-by-step overview of how to use them. To address this need, we have launched a suite of tutorial videos to help our customers better use our products.

Visikol HISTO - Getting Started Tutorial

Confocal Microscopy Tutorial

Light Sheet Microscopy Tutorial

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Visikol and InSphero App Note

Check out Visikol and InSphero's recent application note that features the use of Visikol HISTO-M with InSphero's microtissue models. It was shown that combining Visikol HISTO-M with confocal microscopy allows for a 3-fold increase in the number of cells detected in microtissues.

Check out the full application note here

Visikol and InSphero Application Note

Teaming Up With Young Talent to Disrupt Bio-imaging

Medical imaging has made remarkable leaps and bounds. In today's world, doctors and scientists can generate rich images from a multitude of sources, such as confocal microscopes and MRI machines. With popular programs such as the open-source, powerful ImageJ editor, these images can be further enhanced and even be used to construct accurate three-dimensional models of the original sample. However, despite breakthroughs in virtual reality technology, these 3-D models and image stacks are still viewed in a two-dimensional context: through a monitor. The low usage of inexpensive VR solutions such as Google Cardboard can be attributed to the time and expertise normally required to create a VR compatible model or video clip. To allow for simple integration into VR technology, Visikol teamed up with Somerville High School student Joe Napoli to write a Stereoscopic 3D Projection plugin for ImageJ.

Visikol HISTO Vasculature Viewer

The Stereoscopic 3D Projection plugin provides a straightforward way to create anaglyph or Google Cardboard-compatible VR projections. Written in Java 8, this plugin takes in an image stack as input. After reading in values such as slice spacing from the image stack's metadata, the user can dictate how many degrees the projection will rotate before generating a new frame, how many degrees the final projection will rotate, the start and stop rotation of the projection, and whether the projection is done in anaglyph or for Googe Cardboard. Here, the user can also adjust the slice spacing, if necessary. To generate these projections, the plugin utilizes ImageJ's powerful innate 3D projection function to create projections for the left and right eyes. The right eye projection is rotated an additional five degrees, to account for the spacing between one's eyes. If the projection is in anaglyph format, the left and right eye projections are colored red and blue, respectively, and then merged into one model. For the Google Cardboard format, however, the image stack retains its original color and the left and right eye projections are placed side by side instead of being merged. Regardless of whether the product is formatted for anaglyph or Google Cardboard, the plugin's output can be saved as a video or directly viewed.

Visikol ImageJ Plugin User Interface

 

The Stereoscopic 3D Projection plugin's source, along with screenshots of the plugin in action, can be found on GitHub. The source is also attached to the JAR, located on the plugin's official ImageJ site. The Stereoscopic 3D Projection plugin can be installed through Fiji's updater after adding http://sites.imagej.net/JoeNapoli/ to the list of update sites, or by downloading the JAR file and placing it into ImageJ's plugins folder.

Visikol HISTO Stereoscopic

Governor Chris Christie stops by Visikol headquarters

Chris Christie came by the Visikol lab today to chat about innovation and biotech in NJ. Michael got the chance to introduce the Visikol technology and talk about how we are working to improve bio-imaging. 

CEO and Co-Founder Michael Johnson with Governor Chris Christie 

CEO and Co-Founder Michael Johnson with Governor Chris Christie 

Q2 Quarterly Review

What a great three months for Visikol! I am incredibly proud of the progress that we have made in reaching our milestones and shifting the paradigm of histology from 2D to 3D. Over the last few months we have focused entirely upon improving our Visikol HISTO technology and are looking forward to the second half of the year with many exciting announcements to come. Below you can read about what we have been focusing on for the last three months.

Improved User Experience:

In launching any new product whether it is Airbnb or Visikol HISTO, the user experience is critical as building a product that works in your hands is not enough – The product needs to perform flawlessly in the hands of ALL your users. With Visikol HISTO, we have focused a lot of our time on improving the customer experience so that any researcher regardless of background or skill set can successfully drop the technology into their existing bio-imaging workflow. We have done this through an extensive beta test and partnering with researchers in diverse fields. Through this process, we gained an intimate understanding of how our customers use the Visikol HISTO technology and in Q2 launched the Visikol HISTO protocol builder to help improve the overall customer experience. This web app was built by Nick Crider and addressed our biggest shortcoming with Visikol HISTO which was that customers wanted customized protocols for THEIR specific applications. Moving forward, our focus will be on continually improving the user experience so that customers can more quickly and successfully drop the Visikol HISTO technology into their workflow. In Q3, we will begin launching a suite of tutorial videos on 3D bio-imaging as well as application specific Visikol HISTO kits, image processing plugins and a webinar series.

Build out of Visikol HISTO bio-imaging platform as a service (i.e. HISTOPlex):

One of the long-term goals for Visikol is to develop and launch a 3D bio-imaging service business that can assist pharmaceutical companies with the drug discovery process. Specifically, we plan to develop several bio-imaging assays that will allow pharma companies to extract actionable 3D insights from tissues. We are tentatively calling this platform HISTOPlex and it will be a high throughput system that leverages the Visikol HISTO technique. We began developing this platform in Q1 of 2017 and plan to begin offering it in late 2018 to customers. We have shown that for tissues like spheroids commonly used in drug discovery that we can acquire more accurate and complete information than is acquired today. 

Big Data Analysis:

While we will continue to build on the Visikol HISTO technology and improve the user experience, it is essentially complete. This is an exciting time as we are now able to transition Visikol from focusing on building tools for acquiring data to focusing on HISTOPlex to analyze the data. This is a major shift as most publications and work in the tissue clearing space have focused solely on data acquisition from tissues. With HISTOPlex, we are able to generate hundreds of gigabytes of data from tissues and are beginning to develop a platform for storing this data and extracting features of interest. We see HISTOPlex shifting the paradigm of how tissues are characertized and look forward to accelerating the drug discovery process.

I look forward to updating you on our progress in Q3 - stay tuned. 

Michael 

Michael speaking at Rutgers Univeristy about Visikol. 

Michael speaking at Rutgers Univeristy about Visikol. 

Visikol HISTO Trouble Shooting Guide

We have introduced a trouble shooting guide below that address common problems and feedback that we have received from beta testers and customers. The trouble shooting guide describes the sources of each problem and potential solutions. 

I can’t image past 500-800 µm. Labeling appears uneven, and drops off significantly at this depth.

  • Antibody concentration too high: ring of intense staining near surface, drops off significantly after that.
    • Solution: Reduce antibody concentration, if signal is too weak, incubate in lower concentration for half of time, and then re-incubate in higher concentration.
  • Antibody concentration too low: signal drops off into middle of tissue.
    • Solution: Increase antibody concentration.
  • Optical attenuation due to absorption of photons by upper layers of tissue causes “shadow” to tissues below, even with perfect staining
    • Solution: Increase laser power and gain as depth increases. Caution: increased laser power increases rate of photobleaching, be sure samples contain no air bubbles. Leica SP5 and SP8 can automate laser power and gain corrections. Compare intensity loss to nuclear stain intensity, since nuclear stain diffuses very fast into tissue. Can use this signal to correct for signal loss in image processing.

Intense band of labeled tissue at surface, then significant drop-off afterwards.

My tissue didn’t clear!

  • Plastic Incompatibility
    • Visikol HISTO-2 will degrade polystyrene. For processing tissues with the Visikol HISTO approach we suggest moving away from polystyrene and towards polypropylene and glass where possible. Plastic leaching into your sample may affect the clearing ability of Visikol HISTO.
  •  Incomplete Dehydration/Clearing
    • Most of the time a lack of tissue transparency is simply due to not completely dehydrating a sample. If you use methanol or ethanol for dehydration that has water in it and is not pure, you will not remove all the water from your tissue, resulting in tissue cloudiness. This can also be caused by not sealing the vessels containing your sample when clearing, as Visikol HISTO-2 is hygroscopic. Additionally, not using enough volume of Visikol HISTO-1 and Visikol HISTO-2 for your tissue size can cause inadequate clearing.

For a mouse brain that is not completely clear, we suggest placing the brain back into 7-10 mL of Visikol HISTO-1 for 24 hours, followed by transfer to 25 mL methanol for 2 hours. Then transfer back to 7-10 mL Visikol HISTO-1 for 24 hours, followed by 7-10 mL of fresh Visikol HISTO-2 for 24 hours.

Fluorescent protein quenched

  • To visualize fluorescent protein, samples must be dehydrated using t-Butanol instead of methanol.
  • Keep cleared samples in the dark, and protect your specimens with aluminum foil as fluorescent proteins photobleach rapidly when exposed to ambient light.
  • Do not treat fluorescent protein labeled samples with H2O2 bleaching step; this step will oxidize fluorescent protein and signal will be lost.

My antibody didn’t label the tissue

  • Some antibodies are not compatible with 3D immunolabeling. Validate the specificity of your antibody on small tissue sections before proceeding to larger tissues. Contact us if you have any questions about your specific antibody.
  • Only purchase antibodies that have been validated for use in IHC.
  • Please follow our antibody optimization protocol

Center of tissue appears dark

  • Insufficient antibody concentration
    • Solution: Increase antibody concentration. A range of concentrations should be explored on a small section of tissue prior to scaling to large tissues. 
  • Optical attenuation leads to diminished signal at increasing depths depending on several factors, such as concentration of label bound in upper layers of tissue, level of autofluorescence, type of objective, and laser power. 

Tissue has become yellow

Visikol HISTO will cause tissues to become slightly yellow during the clearing process. To reduce this effect, conduct all tissue processing steps at room temperature. This tissue yellowing will effect the orange color channel (TRITC) the most. 

For the full Visikol HISTO Guidebook click here.

Visikol HISTO Guidebook

 

 

 

Launching the Visikol HISTO Protocol Builder

We have assembled all of our knowledge about using Visikol HISTO with various tissues into an easy-to-use tool that will generate a step-by-step protocol specifically for your tissue, label, and microscope. The Protocol Builder web app allows you to select parameters specific to your goals and delivers a printable and shareable protocol so you will be sure to get the best results from Visikol HISTO. Thanks so much to our beta testers for all the feedback we have incorporated into this tool.

To try out the protocol builder click below. Let us know what you think and if you have any feedback or questions. 

Visikol HISTO Protocol Builder

Updated Visikol HISTO Guidebook

Over the past six months, we have been compiling all the feedback, considerations, and advice about using Visikol HISTO into our comprehensive Visikol HISTO Guidebook. Our new and improved guidebook has detailed descriptions of every consideration that is important to labeling and tissue clearing. To read through our updated guidebook click below. 

Visikol HISTO Guidebook

Introducing Visikol HISTO-M

Seeing Deeper into 3D Tissue Cultures

Microtissues and organoids are too thick to be fully imaged using confocal microscopy. In PBS, only the outer layers of cells may be imaged.  After treatment with Visikol HISTO-M, cells can be imaged on every plane through the center, allowing you to extract more information from your tissue models.

3D Tissue Culture in PBS

Figure 1. Confocal z-stack montage of NCI-2170 Organoids stained with SYTOX green, in PBS. Notice that only outer layer of cells is visible to imaging. 20 um z-step.

Figure 1. Confocal z-stack montage of NCI-2170 Organoids stained with SYTOX green, in PBS. Notice that only outer layer of cells is visible to imaging. 20 um z-step.

3D Tissue Culture with Visikol HISTO-M

Figure 2. Confocal z-stack montage of Visikol MICRO-treated NCI-2170 organoids stained with SYTOX green. The full plane of cells is visible in each image. 20 um z-step.

Figure 2. Confocal z-stack montage of Visikol MICRO-treated NCI-2170 organoids stained with SYTOX green. The full plane of cells is visible in each image. 20 um z-step.