M\u00fcller, a soft-spoken professor from the University of California, Berkeley, was there to speak about a technique to overcome low contrast in electron microscopy images. Low contrast is one of the key problems in using cryo-electron microscopy (cryo-EM) for structural biology, and M\u00fcller had worked to solve it for a decade.<\/p>\n
As the conference concluded, moderator Stephani Otte asked attendees for one wish. M\u00fcller recalls how the first person to answer named the proof-of-concept instrument that M\u00fcller had presented, \u201cand then the next person gets up and says the same thing.\u201d One by one, he says, nearly everyone at the conference said they wanted to use his instrument.<\/p>\n
After a formal peer-review process, CZI (which last year rebranded three labs and its imaging center into a single entity known as Biohub<\/a>) invested in M\u00fcller\u2019s research. The result of that project, a piece of hardware called a laser phase plate (LPP) that improves the contrast of cryo-EM images<\/a>, has now been published (Science 2026, DOI: 10.1126\/science.aeh0665). Researchers say it represents a dramatic improvement in primary data that could bring structural biology to smaller proteins and could bring new capabilities for in situ structural biology<\/a> within reach.<\/p>\n A very bright laser to improve cryo-EM<\/p>\n To image biomolecules like proteins, the microscopes used for cryo-EM shoot a beam of electrons through a sample. The way that the electrons scatter allows researchers to see what\u2019s there. The microscopes let chemists collect enormous detail on materials<\/a>, such as uncrystallizable proteins and complex biological specimens.<\/p>\n But biological substances are almost completely transparent to electrons, so only a small proportion of the electrons in an imaging beam are scattered. Most of the electrons pass through a sample without interacting with it at all. \u201cThat means that a perfect image gives nearly no contrast,\u201d M\u00fcller says. A raw micrograph resembles a blizzard; researchers obtain detail by averaging many images.<\/p>\n Microscopists solved an analogous problem with visible light in the 1930s by inventing phase-contrast microscopy. This approach takes advantage of the phase difference between scattered and unchanged light. A filter plate shifts the phase of unscattered background light back into phase with the scattered light. The resulting interference between the two types of photon turns the invisible phase difference into a detectable difference in brightness.<\/p>\n But it has been much harder to find a material that could be used to make a filter plate for electrons. \u201cIf you put something physical in the microscope, it interacts in a bad way with the electrons themselves: it charges up, it deteriorates, it changes,\u201d says David Agard, the founding scientific director of imaging at Biohub.<\/p>\n M\u00fcller and his colleagues theorized back in 2010 that one might use photons to shift electrons\u2019 phase<\/a> instead (New J. Phys., DOI: 10.1088\/1367-2630\/12\/7\/073011). By shooting a laser at a microscope\u2019s electron beam after it passes through a sample, they calculated, one could alter electrons that had not been scattered by the sample more dramatically than those that had.<\/p>\n The only problem was it would take a laser brighter than anyone had ever built.<\/p>\n M\u00fcller says he thought it should be possible, but he wasn\u2019t absolutely certain. \u201cPhysics calculations have a habit of being off by \u03c0 or 2\u03c0 or something like that, because you screwed up somewhere.\u201d<\/p>\n “Physics calculations have a habit of being off by \u03c0 or 2\u03c0 or something like that, because you screwed up somewhere.”<\/p>\n Holger M\u00fcller, physicist, University of California, Berkeley<\/p>\n Share To make the exceptionally bright laser, M\u00fcller and his lab developed an optical resonator\u2014two perfectly aligned, highly smooth spherical mirrors. When light from a 12 W laser enters, it bounces back and forth between the mirrors, picking up more energy from the incoming laser with each reflection. Eventually, it reaches a very bright equilibrium. \u201cThe laser phase plate has the highest continuous light intensity in the solar system, outside of the inner regions of the sun,\u201d M\u00fcller says.<\/p>\n The physicists also engineered feedback loops to keep the mirrors aligned as they heat up under the illumination, which is equivalent to the output of three power plants per square millimeter. After more than a decade of work building an LPP into an older electron microscope, they showed it could shift the phase of the electron beam<\/a> and hold steady for long enough to capture a structure (Rev. Sci. Instr. 2021, DOI: 10.1063\/5.0045496).<\/p>\n This study was the proof of principle that M\u00fcller presented at the CZI conference. To find out if the LPP could improve on the state of the art in microscopy, he needed to install it in an up-to-date microscope.<\/p>\n Laser-on cryoEM captures smaller proteins with greater clarity<\/p>\n With a Biohub grant, M\u00fcller and his team installed an LPP into a modern electron microscope adjusted to leave space for the extra hardware. They found it delivered much higher-resolution structures of the smallest protein they tested.<\/p>\n Joachim Frank<\/a> at Columbia University, who received the 2017 Nobel Prize in Chemistry <\/a>for his role in inventing cryo-EM, says in an email to C&EN that the advance is \u201chighly significant as it opens up the whole technology for much smaller biological molecules.\u201d<\/p>\n Capturing the shape of purified small proteins, and the way they interact with ligands, could be a boon for the pharmaceutical industry. But what especially excites many in the structural biology<\/a> community is the prospect of identifying small proteins in heterogeneous mixtures\u2014like images from cell samples, where researchers currently can pick out just a few large features.<\/p>\n Improving \u201cthe weak contrast that you get in single-particle [cryo-EM] is even more crucial for tomography, because you just don\u2019t know what you\u2019re looking at without more contrast,\u201d says Anthony W. P. Fitzpatrick<\/a>, a Columbia University biophysicist who received a grant from CZI for a related approach to phase shifting, manipulating an electron beam with a bright high-speed laser pulse<\/a>. (Fitzpatrick recently published on the approach<\/a> but has yet to install it into a microscope.) He adds, \u201cThe challenge will be the price. The price will be astronomical.\u201d<\/p>\n Postdoctoral researcher Jessie Zhang prepares to install a laser phase plate in a cryo-electron microscope. She works under a HEPA filter; because the laser is so bright, the tiniest grain of dust in the spherical mirrors used to amplify it can incinerate and ruin the mirrors. \u201cWe became so paranoid to keep the mirrors clean,\u201d Holger M\u00fcller says.<\/p>\n Credit: According to Bronwyn Lucas<\/a>, a Berkeley biophysicist who has been working with M\u00fcller to develop tomography approaches using the LPP, the tool is \u201cmassively expanding the\u00a0proportion of the proteome that can be captured in cells.\u201d<\/p>\n In a companion preprint, published before peer review, a group of researchers at Biohub announce that they have built a second LPP-enabled microscope<\/a> (bioRxiv 2026, DOI: 10.64898\/2026.06.05.730245). This one features a different LPP design, called a dual phase plate, which uses two crossed laser beams that demand half the intensity and therefore less-perfect mirrors. M\u00fcller and his team just published the theory supporting the dual phase plate<\/a> (Nat. Commun., DOI: 10.1038\/s41467-026-74060-6).<\/p>\n According to Agard, who is a cocorresponding author on the Biohub preprint showing that the design works, \u201cOne of the referees for that paper had said, \u2018Oh, nobody will ever be able to build this, it’s too challenging, it’s too complex.\u2019\u201d But they did. Besides showing that phase-contrast cryo-EM can work in someone else\u2019s hands, the preprint also includes a proof-of-concept image of a slice of a bacterial cell.<\/p>\n Research teams at Berkeley and Biohub are continuing to push the two LPP-equipped microscopes from demonstration projects toward genuine data collection. M\u00fcller says he\u2019s focused on collecting images in sharper focus.<\/p>\n While there\u2019s a lot more work to do, Agard says, he hopes soon that \u201cwe\u2019ll be able to really start doing structural cell biology at scale, which for us has been a dream.\u201d<\/p>\n \u201cThe big hope here is that cryo-electron tomography would have a similar resolution revolution to what single-particle cryo-EM had,\u201d Fitzpatrick says. It hasn\u2019t happened yet. But he adds, \u201cFor tomography to have that big jump\u00a0.\u00a0.\u00a0. the primary data itself has got to get better.\u201d<\/p>\n
\n <\/a><\/p>\n![\"A](\"https:\/\/www.newsbeep.com\/ie\/wp-content\/uploads\/2026\/06\/EMBARGOED-short-feature---phase-contrast-for-electron-microscopy---488259.webp\")
\n A person in a cleanroom suit works hunched under a HEPA filter inside the chassis of an electron microscope, which features lots of hardware.<\/p>\n
\n Petar Petrov<\/p>\n
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