Forum for Science, Industry and Business

Sponsored by:     3M 
Search our Site:

 

New technique efficiently turns antibodies into highly tuned 'nanobodies'

03.11.2014

Antibodies, in charge of recognizing and homing in on molecular targets, are among the most useful tools in biology and medicine. Nanobodies – antibodies' tiny cousins – can do the same tasks, for example marking molecules for research or flagging diseased cells for destruction. But, thanks to their comparative simplicity nanobodies offer the tantalizing prospect of being much easier to produce.

Unfortunately, their promise hasn't been fully realized, because scientists have lacked an efficient way of identifying the nanobodies most closely tuned to their targets. However, a new system, developed by researchers at Rockefeller University and their collaborators and described today in Nature Methods, promises to make nanobodies dramatically more accessible for all kinds of research.

Antibodies are defensive proteins deployed by the immune system to identify and neutralize invaders. But their power can be harnessed in other ways as well, and they are used in biology and medicine for visualizing cellular processes, attacking diseased cells and delivering specific molecules to specific places. Like their larger cousins, nanobodies can also be used for these tasks but their small size makes nanobodies much easier to grow in bacterial factories. They can also access hard to reach places that may be off limits to larger molecules.

"Nanobodies have tremendous potential as versatile and accessible alternatives to conventional antibodies, but unfortunately current techniques present a bottleneck to meeting the demand for them," says study author Michael Rout, head of the Laboratory of Cellular and Structural Biology at Rockefeller. "We hope that our system will make high-affinity nanobodies more available, and open up many new possible uses for them."

... more about:
»GFP »Laboratory »Rockefeller »immune »proteins »sequence

In their first studies, the team generated high-affinity antibodies, those that are capable of most precisely binding to their targets, directed against two fluorescent proteins that biologists often use as markers to visualize activity within cells: GFP and mCherry. Their new system, like existing ones for generating antibodies, begins with an animal, in this case llamas housed in a facility in Connecticut.

Llamas were chosen because the antibody variants they produce are easily modified to generate nanobodies, which are only one-tenth the weight of a regular antibody. The llamas were immunized with GFP and mCherry, prompting their immune systems to generate antibodies against these foreign proteins.

"The key was to figure out a relatively fast way of determining the genetic sequences of the antibodies that bind to the targets with the greatest affinity. Up until now obtaining these high-affinity sequences has been something of a holy grail," says Brian Chait, Camille and Henry Dreyfus Professor and head of the Laboratory of Mass Spectrometry and Gaseous Ion Chemistry at Rockefeller. "Once those sequences are obtained, it's easy to engineer bacteria to mass produce the antibodies."

The researchers, led by graduate student Peter Fridy and postdoc Yinyin Li, started by making antibody sequence databases from RNA isolated out of antibody-producing cells in the llamas' bone marrow. Next, they picked out the tightest binding GFP and mCherry antibodies from blood samples from the same llamas, and chemically cut these into smaller pieces, keeping only the antigen-binding section to create nanobodies.

They then determined partial sequences of the amino acids that made up the protein of the nanobodies with a technique known as mass spectrometry. Using a computer algorithm called "llama magic," developed by David Fenyö and Sarah Keegan of New York University School of Medicine, they matched up the composition of the highest affinity nanobodies with the original RNA sequence. With this sequence, they could engineer bacteria to mass produce the nanobodies before putting them to use in experiments.

Antibodies are often used to isolate a particular structure within a cell so scientists can remove and examine it, and the team did just that with their new nanobodies. They purified various cellular structures tagged with GFP or mCherry, and also visualized these structures in situ.

All in all, their procedure generated 25 types of nanobodies capable of precisely targeting GFP and six for mCherry, a far more diverse set of high affinity nanobodies than is typically possible with conventional techniques.

This abundance opens up new options. Scientists can select only the best ones, eliminating nanobodies that by chance cross-react with other molecules, or string together two nanobodies that attach to different spots on the same target molecule to generate a super-high-affinity dimer, exactly as the researchers demonstrated for the GFP nanobodies. This super-high-affinity could be a powerful feature when delivering therapeutic or diagnostic molecules because it would lower the required dosage, and so reduce unwanted side effects.

"Given that we can now readily identify suites of high affinity nanobodies, the future for them as research tools, diagnostics and therapeutics looks bright," says Rout.

Zach Veilleux | EurekAlert!
Further information:
http://www.rockefeller.edu

Further reports about: GFP Laboratory Rockefeller immune proteins sequence

More articles from Life Sciences:

nachricht Multi-institutional collaboration uncovers how molecular machines assemble
02.12.2016 | Salk Institute

nachricht Fertilized egg cells trigger and monitor loss of sperm’s epigenetic memory
02.12.2016 | IMBA - Institut für Molekulare Biotechnologie der Österreichischen Akademie der Wissenschaften GmbH

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

Die letzten 5 Focus-News des innovations-reports im Überblick:

Im Focus: Novel silicon etching technique crafts 3-D gradient refractive index micro-optics

A multi-institutional research collaboration has created a novel approach for fabricating three-dimensional micro-optics through the shape-defined formation of porous silicon (PSi), with broad impacts in integrated optoelectronics, imaging, and photovoltaics.

Working with colleagues at Stanford and The Dow Chemical Company, researchers at the University of Illinois at Urbana-Champaign fabricated 3-D birefringent...

Im Focus: Quantum Particles Form Droplets

In experiments with magnetic atoms conducted at extremely low temperatures, scientists have demonstrated a unique phase of matter: The atoms form a new type of quantum liquid or quantum droplet state. These so called quantum droplets may preserve their form in absence of external confinement because of quantum effects. The joint team of experimental physicists from Innsbruck and theoretical physicists from Hannover report on their findings in the journal Physical Review X.

“Our Quantum droplets are in the gas phase but they still drop like a rock,” explains experimental physicist Francesca Ferlaino when talking about the...

Im Focus: MADMAX: Max Planck Institute for Physics takes up axion research

The Max Planck Institute for Physics (MPP) is opening up a new research field. A workshop from November 21 - 22, 2016 will mark the start of activities for an innovative axion experiment. Axions are still only purely hypothetical particles. Their detection could solve two fundamental problems in particle physics: What dark matter consists of and why it has not yet been possible to directly observe a CP violation for the strong interaction.

The “MADMAX” project is the MPP’s commitment to axion research. Axions are so far only a theoretical prediction and are difficult to detect: on the one hand,...

Im Focus: Molecules change shape when wet

Broadband rotational spectroscopy unravels structural reshaping of isolated molecules in the gas phase to accommodate water

In two recent publications in the Journal of Chemical Physics and in the Journal of Physical Chemistry Letters, researchers around Melanie Schnell from the Max...

Im Focus: Fraunhofer ISE Develops Highly Compact, High Frequency DC/DC Converter for Aviation

The efficiency of power electronic systems is not solely dependent on electrical efficiency but also on weight, for example, in mobile systems. When the weight of relevant components and devices in airplanes, for instance, is reduced, fuel savings can be achieved and correspondingly greenhouse gas emissions decreased. New materials and components based on gallium nitride (GaN) can help to reduce weight and increase the efficiency. With these new materials, power electronic switches can be operated at higher switching frequency, resulting in higher power density and lower material costs.

Researchers at the Fraunhofer Institute for Solar Energy Systems ISE together with partners have investigated how these materials can be used to make power...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

Event News

ICTM Conference 2017: Production technology for turbomachine manufacturing of the future

16.11.2016 | Event News

Innovation Day Laser Technology – Laser Additive Manufacturing

01.11.2016 | Event News

#IC2S2: When Social Science meets Computer Science - GESIS will host the IC2S2 conference 2017

14.10.2016 | Event News

 
Latest News

UTSA study describes new minimally invasive device to treat cancer and other illnesses

02.12.2016 | Medical Engineering

Plasma-zapping process could yield trans fat-free soybean oil product

02.12.2016 | Agricultural and Forestry Science

What do Netflix, Google and planetary systems have in common?

02.12.2016 | Physics and Astronomy

VideoLinks
B2B-VideoLinks
More VideoLinks >>>