With chicken eggs and household supplies, undergraduates blaze a path toward low-cost antiviral

With chicken eggs and household supplies, undergraduates blaze a path toward low-cost antiviral

Posted on April 11th, 2022

Using household items, Stanford students have developed a way to make affordable nasal drops with the potential to slow the spread of viruses like COVID-19.

Re-post from Stanford Medicine | April 8, 2022

Stanford researchers, including six undergraduate students, have created an inexpensive method for making nasal drops that could stem the spread of viruses such as COVID-19. The approach could be particularly useful for people living in low-resource countries.

Using chicken eggs and household items, they devised a way to extract and purify antibodies present in yolks — called immunoglobulin Y (IgY) — that are proven to be safe and may prevent or treat many infectious diseases. The students identified a way to purify the IgY without a commercial laboratory or expensive chemicals and equipment, as long as electricity and a freezer are available.

“IgY purified from eggs of immunized hens could be a tool to slow down the spread of airborne viruses,” said Daria Mochly-Rosen, PhD, the George D. Smith Professor of Translational Medicine, who oversaw the research team.

“Using IgY is not to replace vaccines, but complement them,” she added. “Only about 10% of people living in low-income countries are immunized against SARS-CoV-2 in the third year of the pandemic. This is not good enough.”

The study was published Feb. 26 in the Journal of Global Health. Mochly-Rosen, president and founder of Stanford’s SPARK Global program, which helps translate biomedical research into clinical applications through university-industry partnerships, was the senior author. The students were co-authors.

Although the nasal drops would likely be effective for only a few hours, Mochly-Rosen noted, they may provide protection when people enter a crowded space, such as an airplane or classroom. Moreover, as a virus changes or a new one appears, hens could be immunized with the new variant protein, generating new IgYs, which can then be purified and distributed faster than a new vaccine, she said.

The researchers chose nasal drops as the delivery system based on evidence that COVID-19 transmission can be reduced at points of entry into the body, which include the nose. The IgY antibodies work by binding to the virus and preventing its access to the receptors in the nasal passage.

An antibody factory
Chicken eggs naturally contain antibodies, which protect newly hatched chicks from diseases. Hens will produce eggs that contain antibodies to a new virus if the hens are injected with proteins (antigens) to that virus. It takes about three weeks after injection for effective antibodies to show up in the eggs.

Mochly-Rosen started researching antibodies against SARS-CoV-2 from the eggs of immunized hens in mid-2020. A trial in Australia found the nasal drops to be safe.

In June 2021, Mochly-Rosen and her team followed that up with a review paper published in Frontiers in Immunology detailing the many potential uses of chicken IgY to safely detect, prevent and treat animal and human infections and infectious diseases, including hepatitis B, Zika virus and dengue fever.

But all of that led to a burning question for Mochly-Rosen: Was there a low-cost way to extract the IgY antibodies from eggs, one that could be used in areas without a commercial laboratory?

Mochly-Rosen posed the question to a group of Stanford freshmen, who signed up for the research through Stanford Students in Biodesign and Biopharma, a club for those interested in medical technology.

“All of these students were smart cookies,” she said. “We met [online] every Monday at 9 p.m. Pacific time … and discussed how to crack eggs, how to isolate the antibodies using household ingredients and how to adapt household equipment.”

Because their research mostly was conducted last year, when COVID-19 restrictions limited on-campus gatherings, the students worked in their homes and dorm rooms. They used ordinary household materials, such as baking soda and vinegar, and kitchen equipment, such as a food processor.

The research “was an incredible opportunity, and it was in line with my focus on equity and accessibility for protection against COVID around the world,” Allison Jia said. She and a fellow researcher “were holed up in her dorm room cracking eggs to find the best way to extract the yolk. We accidentally stunk up the women’s bathroom getting rid of the eggs.”

They tested the pH levels of various ingredients, such as grapefruit and lemon juice, to find the best substitute for commercial chemicals. The team chose vinegar, partly because it’s more widely available than fresh fruit in many places, said Jia, now a sophomore.

Other students were tasked with finding household products that would spin at high speed to simulate a centrifuge. They tried an electric razor and a blender before settling on a food processor modified to hold test tubes, Mochly-Rosen said.

How to make antiviral nose drops
The students worked with eggs purchased from grocery stores, online grocery delivery services or other sources, so the eggs lacked antibodies to viruses such as SARS-CoV-2. However, the eggs contained other antibodies, which the students were able to isolate. The isolation process is the same for any antibody.

Here, in brief, is the recipe they devised: Separate the yolk from the white, then dilute the egg yolk in water. Add vinegar, shake gently, then freeze until solid. Thaw and filter the mixture to remove the fat. Add salt and place the solution in test tubes in a commercial or makeshift centrifuge, where gravitational forces cause soft beads to form at the bottom of the tubes. Dissolve the beads in water, neutralize them with baking soda and place the solution in a dropper.

Lab analyses confirmed the protocols worked as well as, if not better than, commercial processes, Mochly-Rosen said. Under the students’ method, each egg produced about 90 milligrams of IgY antibodies, or about 25 doses. The purified IgY solution is stable at room temperature for up to two weeks and for much longer if prepared under sterile conditions and refrigerated.

The researchers also developed a kit that includes everything needed for the entire process — including an egg yolk separator, gloves, bottles, salt, pH strips and pipettes. There are two kit versions: one with a makeshift centrifuge (a food processor with instructions to turn it into a centrifuge using a 3D printer) and one without (for labs that have a commercial centrifuge).

All of the kit materials, except the food processor, fit into a 15-inch box.

A plan for distribution
According to the researchers, an industrial or academic laboratory could produce a COVID-19 antigen or an antigen from another virus and send it to chicken farmers to inject into their hens. The farmers then could distribute the antibody-egg-laying hens or their eggs.

Mochly-Rosen hopes food processor manufacturers would be willing make an improvised centrifuge at cost for rural areas that don’t have access to commercial centrifuges.

Their price analysis suggests that one dose using the kit would cost as little as 20 cents compared with $5.40 in a commercial setting.

“This could be an effective and cheap solution,” Mochly-Rosen said. “If we’re trying to address an epidemic or pandemic properly, we need to make sure a solution is available fast and everywhere in the world.”

Other Stanford co-authors to the study are Ravinder Pamnani, instructor at the Stanford Byers Center for Biodesign; undergraduates Carrie Chen, Anna Hudson, Caitlin Kunchur, Andrew Song and Edward Tran; graduate students Chris Fisher Davide Zanchi, PhD; life science researcher Lucia Lee; and former postdoctoral scholar Ana Koperniku, PhD; Stephen Kargotich, executive director of SPARK Global; and Mary Romeo, science and communications writer at SPARK Global.

The research was funded by SPARK at Stanford Translational Research Program and the Stanford Byers Center for Biodesign.