A New Platform for Protein Detection

AmberGen’s Photocleavable Mass Tag (PCMT™) Technology

At the heart of AmberGen’s MALDI-IHC platform are its patented photocleavable mass-tags (PCMTs). These mass-tags act as barcodes, which when detected by a MALDI Mass Spectrometer, enable simultaneous imaging of hundreds of targeted intact proteins in a tissue or dispersion of cells. This eliminates the need for iterative cycling required for some high-plex fluorescence imaging platforms.

A key feature of these novel PCMTs is the ability to readily conjugate them to antibodies or other molecular probes in a 1-step high-yield chemical reaction. The resulting Miralys™ antibody probes consist of peptide mass-tags covalently attached through patented PC-Linkers to an antibody with an optional fluorophore for multimodal fluorescence and MALDI imaging on the same sample. Miralys™ antibody probe staining of tissues is accomplished using standard manual or automated immunohistology protocols and equipment. After staining, the tissue is pre-UV photocleaved prior to matrix deposition and MALDI-MSI. Importantly, the photocleaved peptide mass-tag possesses a positively charged amino terminated ion which facilitates MS detection. Detection of each mass-tag ion produces a discrete peak at known m/z in the mass spectrum allowing reconstruction of 2D images of the spatial distribution of the targeted protein in the tissue slice.

technology infographic

Key features of AmberGen’s photocleavable mass-tags (PCMTs) and Miralys™ imaging include: Top: 1-step conjugation to probes such as antibodies; Middle: PC-Linkers which upon photocleavage release peptide-mass tags with a terminal positively charged amino group; Bottom: Mass-tag coding of targets to produce discrete peaks at known m/z.

Some key advantages of Miralys Imaging include:

  • HyperPlex Panels: Since hundreds of PCMTs can be used together, many ultra-high-plex panels can be created. Since the mass tags peaks don’t overlap in the mass spectrum, the panels can be combined, creating even higher plex panels. Miralys panels can be configured from Miralys standard probes or provided pre-mixed and pre-optimized. Unlike high-plex fluorescence imaging methods, Miralys imaging does not suffer from autofluorescence, fluorophore bleaching, or spectral overlap.
  • Spatial Multiomic: Since it is based on MSI, both unlabeled small molecules such as metabolites and targeted macromolecules such as proteins can be imaged in the same tissue sample, on the same instrument. For example, both small molecule drugs and their protein targets can be imaged on a single sample. Additional workflows allow imaging of glycans, extracellular matrix proteins and nucleic acids heralding a new dimension of spatial multiomic imaging.
  • Spatial Multimodal: Dual-labeled fluorescence and mass-tag Miralys antibodies allow both fluorescence and MS imaging on the same sample using the same probes. This enables high spatial cellular resolution for selected targets followed by lower resolution rapid MSI scans. Combined fluorescence/MS using the same probes means the fluorescence and MALDI-images can be easily co-registered to provide increased information from the tissue.
  • Simultaneous Staining, Stability and Reimaging: All targeted biomarkers in a tissue are stained simultaneously. Stained slides can be stored for months for later imaging. This enables batch staining of all slides for studies with many samples, reducing experimental variability. Slides that have already undergone MALDI imaging can also be recycled with the same or new Miralys panels.
  • Rapid Whole-Slide Imaging: Hundreds of PCMTs can be imaged simultaneously, meaning that hundreds of proteins can be imaged in a single scan. This vastly surpasses old paradigms for both plexity and speed. For example, 150+ proteins can be imaged as fast as 45 minutes on a 1 cm2 sample or a tissue microarray (TMA) with 100 1-mm2 cores on a standard slide scanned in less than 1 hour.

“Innovation lies at the heart of invention. This has led AmberGen to develop more powerful approaches and products enabling biomedical researchers to uncover the molecular basis of diseases and to develop new therapeutic solutions.”

Ken Rothschild, AmberGen Founder