Overview

The lack of any significant mass spectral overlap (no need for compensation) and auto-fluorescence background makes this technology uniquely suited for unravelling high dimensional functional and phenotypic correlations at the single cell level, accelerating biomarker discovery and drug screening.

image-flow-hyperion-helios

Our BRC Flow Cytometry platform was the first to successfully establish a Mass Cytometry service in the UK and now operates two mass cytometry technology platforms, a Fluidigm HeliosTM (see above right) for suspension cell work and a Fluidigm HyperionTM, imaging mass cytometer (see above left). Imaging mass cytometry uses the same basic principles of established suspension cell mass cytometry to analyze FFPE and frozen tissue sections.

The core offers a full service including custom antibody labelling, panel design, acquisition, troubleshooting and high dimensional data analysis. Data analysisis supported by a dedicated bioinformatician for custom workflows and a highly experienced team of mass cytometry specialists. In addition the BRC Flow Cytometry Platform offers training courses in mass cytometry and high dimensional data analysis.

With currently 38 stable lanthanide isotopes for antibody tagging and more than 10 markers for barcoding and cellular identity, we are able to perform deep phenotyping on clinical and research samples from internal and external clients. Staining protocols and workflows are similar to traditional fluorescent Flow Cytometry as shown in the figure below (modified from Fluidigm).

image-flow-tech-CYTOF-diagram

The figure above gives a brief summary on how liquid mass cytometry works. A single cell suspension of fixed cells labelled with metal-tagged antibodies/probes (A) is introduced into a nebulizer at a rate of ~300 cells/sec creating an aerosol (B) and then transported towards an argon plasma (C) where the cells are vaporized, atomized and ionized.  After selective removal of  low mass ions in the quadrupole  (D),  the ion cloud containing the isotopic metal tags used for labelling enters the TOF chamber where probes are separated by time of flight based on their mass to charge ratio as they accelerate towards the detector (E).  The time-resolved detector measures a mass spectrum that corresponding to the identity and quantity of each isotopic probe on a per-cell basis (F).  Data is saved in FCS format (G) and can be analysed using third party software (H).

image-flow-hyperion

The figure above gives a brief summary on how imaging mass cytometry works.  A tissue sample containing fixed tissue/cells labelled with metal-tagged antibodies/probes (A) is introduced via autostage into the Hyperion Imaging analyser (B) and ablated with a high power pulsed UV laser focused on a 1mm spot size and operating at 100-200 Hz, resulting in plumes of ablated tissue atoms in an inert carrier gas (C) which are then transported towards an argon plasma of the Helios mass cytometer (D) where the arriving atoms are ionized. After selective removal of  low mass ions in the quadrupole (D),  the ion cloud containing the isotopic metal tags used for labelling enters the TOF chamber where probes are separated by time of flight based on their mass to charge ratio as they accelerate towards the detector (E). The time-resolved detector measures a mass spectrum corresponding to the identity and quantity of each isotopic probe on a per-cell basis (E). Data is saved in MCD and TIFF format and can be analysed using third party software reconstructing the tissue architecture alongside high dimensional phenotyping (F).

Antibody conjugation service

The BRC Flow Cytometry platform offers an antibody conjugation service for mass cytometry, bookable via ilabs. The facility provides the conjugation kit for tagging with lanthanide metals (mass range 141-176), all labelling consumables/reagents, manpower and expertise. The customer is requested to provide the antibody raw material. We require at least 100 mg of a protein free IgG solution, the facility will not purify or conjugate antibodies provided in BSA/gelatin containing buffer. We will return the metal conjugated antibody in protein stabilizing solution alongside a quality control data sheet with proof of concentration and successful metal conjugation (biological testing is to be done at customer site). External customers will discuss their project and provide a PO for the agreed project cost before the start of any work.

Further information

All mass cytometry platforms and services are operated by fully trained staff only.

Instruments and services are booked via iLabs. New users must register with iLabs (PDF 235Kb) and complete a Flow Core Registration Form (23kb) prior to first use. To reserve time on our mass cytometers, please make a booking on the instrument calendar. The facility staff will review your request and confirm (or reject) your booking, you will receive a notification via ilabs. Before the first mass cytometry experiment users are required to book a consultation session by contacting us via brcflowcore@gstt.nhs.uk.

To join the UK CyTOF user group please visit us at www.cytof.uk