From left: Claire Fernandes, Arvin Persaud, Christina Guzzo, Deepa Chaphekar, and Aiman Farheen

1 December 2025

By Aideen Teeling

Researchers at the University of Toronto Scarborough (UTSC) in the Guzzo Lab are redefining what we know about human immunodeficiency virus (HIV) biology by developing an emerging technique called flow virometry, which allows them to characterize human proteins on the viral surface.

HIV, which causes acquired immunodeficiency syndrome (AIDS), has been studied extensively since its discovery in 1983. In this time, scientists have uncovered how the virus transmits and establishes infection, and new treatment strategies to manage this chronic infection.

One area of ongoing research explores the ways in which the virus interacts with cells in our bodies, and which proteins mediate these interactions.

Through decades of research, scientists have characterized the viral proteins which scatter the surface and inner core of HIV and the roles these proteins play during infection. However, it has been largely thought that it was mainly viral proteins which facilitated the interactions with and infection of new cells, until now.

A key question for Christina Guzzo, an associate professor in the department of biological sciences at UTSC, is what roles do human proteins play in virus biology and infection. While human proteins had been identified on the surface of HIV before, Guzzo wanted to explore the possibility that these proteins were biologically active and were being hijacked by the virus, and even further, if the proteins had different functions when on a viral particle compared to a human cell.

“For a long time, we’ve been really focused on viral proteins […], but what other proteins might the virus also have on it? It’s not just about immune proteins that are present on the target cells, but also immune proteins can decorate viruses too — I’m interested in this interplay,” she says.

To study these questions, she homed in on an underutilized technique called flow virometry, which uses fluorescently-tagged antibodies to identify proteins on viral particles. Optimizing this technique became a primary focus in the initial years of starting her lab.

Guzzo uses the metaphor of a single-file parade of soldiers to describe flow virometry, where each person is a viral particle, and each one’s hat, coat, gloves and boots represent proteins on the surface, some of which are encoded by the virus genome, and others which are stolen from the host cells that the virus infects.

Flow virometry is a method of rapidly evaluating the soldiers in the line, one-at-a-time and seeing what they’re wearing and how it can be useful to them, she explains. The technique also enables researchers to zoom out and consider the entire parade of soldiers as a group, quantifying the number of viruses with a particular protein of interest in just a few seconds.

According to Guzzo, what sets flow virometry apart from other traditional virology techniques is “the power of analyzing so many [virus] particles in such a short time, so being able to parade 100,000 virus particles […] within just 30 seconds of acquisition”.

Guzzo was first intrigued by the concept of flow virometry during her postdoctoral fellowship at the National Institutes of Health (NIH), where she engaged with colleagues who initially coined the term and technical approach while also working on HIV studies.

During her time at the NIH, Guzzo published a study in Science Immunology where she discovered that a host protein, integrin α4β7, was present on HIV particles and helped direct viruses to the gut, a key site for HIV infection. This work showed how host proteins can play an active role in propagating infection, and redefined host-pathogen dynamics at the cellular level for HIV.

“I realized that one single human protein on a virus can really change where a virus goes,” she says. “Human proteins do things on viruses, and they’re not even part of the viral genome! Sneaky viruses find a way to pick up additional human proteins and hijack them from our cells for their own use.”

Guzzo knew there could be other human proteins that HIV co-opts to maximize infection and wondered whether these human proteins could provide insight into where the virus is replicating in the body. While the prevailing opinion in the field was that all HIV particles look the same regardless of the tissue it replicates in, she saw an application for flow virometry to identify a molecular “fingerprint” of host proteins on the virus, which could provide insight into what types of cells HIV is replicating in, with applications to develop more targeted treatments.

“If you use a new approach to ask an old question, you are probably going to get new answers,” says Guzzo, who is also an associate professor cross-appointed in the department of immunology at the Temerty Faculty of Medicine.

When she came to UTSC to start her own research group in 2017, Guzzo was on a mission to refine and optimize flow virometry to better understand the host proteins on the surface of HIV and other viruses.

“I just saw there was an opportunity in the field to develop new tools, like flow virometry, that could be broadly applicable across the HIV field and virology in general,” says Guzzo, who also sits on the steering committee for the Emerging & Pandemic Infections Consortium.

“It was definitely quite an uphill battle in the beginning,” says Guzzo, explaining that there no papers to reference at the time regarding the quantitative numbers of host proteins on the surface of viral particles, nor any established protocols for performing flow virometry on native, unaltered virus particles. Her team tested nearly every sample in their freezer to establish benchmarks for quantifying human proteins on HIV, with a central goal to ensure their techniques were reliable and reproducible.

Now, in just seven years, the Guzzo lab has pioneered flow virometry so that users can achieve reproducible results between different labs and using different instruments. In an article published earlier this year, her team outlines a rigorous set of guidelines and standardized workflows to enable researchers to use flow virometry to reliably quantify the number of host and viral proteins on the surface of a single particle, or a group of virus particles, with a unique advantage to perform these analyses on viruses in their near-native state.

Not only is the approach less labor intensive, more sensitive, and more user-friendly than other techniques,  her lab’s optimization of flow virometry has also eliminated many sources of background noise, tremendously enhancing the quality of the data.

“We spent a lot of time in the beginning just making sure it was reliable,” says Guzzo, who is the Canada Research Chair in Single Virus Particle Analyses. She adds it was important to establish specific protocols to ensure the data “stand the test of time.”

Guzzo’s vision for HIV research in her lab is to use their refined flow virometry method to uncover new insights into which host proteins help the virus become more effective at transmitting and establishing infection in a new host, which may pave the way for new treatment strategies.

Currently, her team is also working to develop a “multiplex” approach that allows multiple proteins to be detected at the same time on a single virus particle, and additional methods to target proteins inside the virus. In addition, the lab is now applying flow virometry to study influenza viruses, which share similar structures and pathways of infection as HIV. Guzzo says the development of new techniques in HIV research, like flow virometry, is important because it can lead to faster outcomes when applied to other viruses like influenza. This is especially important for the deployment of rapid research pipelines during health emergencies, as was seen in the COVID-19 pandemic.

The Guzzo lab’s work on flow virometry has been central in redefining the role of host proteins on HIV and how we think about viruses. Discovering biologically active host proteins on viruses “really challenged the dogma, or at least the perception of what virus particles look like and how they behave, and it has challenged us to expand our perception of what viruses look like, and the realm of biological activity they can possess,” she says.

“That’s the craft — learning how to make your science useful and leaving your mark in the field so that people can pick it up and build on it. It’s the most important thing we can do as scientists.”