Imaging Cytometry

AmnisⓇ ImageStreamX Mk II Imaging Cytometer

The ImageStream®X Mk II Imaging Flow Cytometer combines the speed, sensitivity, and phenotyping abilities of flow cytometry with the detailed imagery and functional insights of microscopy. In addition to the fluorescence intensity values that are typically provided by flow cytometric measurements, the ImageStream also captures a 12-channel image of each cell, including brightfield, side scatter, and fluorescence, through camera-based detection of a single-cell suspension. Unique time-delay integration of signal attributes signals measured on a pixel-by-pixel basis among all 12 channels to each individual particle, facilitating extremely high-sensitivity measurements. The throughput afforded by the ImageStream’s flow cytometry-style measurement system provides statistical significance of large data sets and allows signal quantification and spatial information of rare populations. This unique combination enables a broad range of applications that would be impossible using either technique alone.

The CSCI Flow Cytometry ImageStreamX Mk II is fully-configured with the following features:

Four excitation lasers for fluorescence-based measurements

  • 405 nm
  • 488 nm
  • 561 nm
  • 642 nm

12 imaging channels across two proprietary CCD cameras

  • Brightfield
  • Side scatter
  • 10 fluorescence channels

Three objective lenses:

  • 60X
  • 40X
  • 20X

You can find the ImageStream optical configuration here.

AutoSampler for unattended 96-well plate acquisition. This sophisticated system provides the following benefits:

  • In-line bubble detection for unattended acquisition
  • Automatic process logging and error notification
  • Automatic sample probe rinse between samples and <0.5% carryover
  • Automatic sample resuspension via aspiration

The Extended Depth of Field (EDF) module keeps depth of cells in focus without loss of sensitivity using Wavefront Coding™ technology from CDM Optics, which is a combination of specialized optics and unique image processing algorithms, to project more structures within the cell into one crisp plane of focus.

The ImageStreamX Mark II is engineered with a variety of sophisticated features that ensure robust performance and facilitate easy-of-use, allowing the machine to be operated by end users after basic training:

  • Automated startup and shutdown procedures, including decontamination
  • On-board fluidics and level monitoring
  • Automated calibration and quality control which incorporates on-board SpeedBeadsⓇ and motorized optics to ensure consistent laser alignment. The automated calibration and QC perform a variety of sophisticated tests to ensure that the machine is operating at peak performance before sample acquisition.
  • Automatic mixing of sample before acquisition
  • Sample probe rinse between samples to prevent carryover (<0.5%)
  • Sample return feature to facilitate virtually 100% sample utilization

Data analysis performed using IDEAS software, which is a powerful platform that incorporates both wizard-based and customized workflows to facilitate data analysis from any application.

Typical applications include but are not limited to the following:

  • Cell Signaling (translocation)
  • Internalization and phagocytosis
  • Intracellular co-localization
  • Shape change and chemotaxis
  • Cell-cell interaction
  • Cell death and autophagy
  • Cell cycle and mitosis
  • DNA damage
  • Stem cell biology (differentiation)
  • Microbiology
  • Parasitology
  • Exosome internalization
  • Microparticle quantitation

IMPORTANT: Because the ImageStreamX MkII was purchased with funds awarded through the NIH S10 Shared Instrumentation Grant program (Award Number S10OD026845), publications utilizing data acquired with the ImageStream must acknowledge this grant. Please use the following language in publications in which you have used the ImageStream for your experiments:

Research reported in this publication using the ImageStreamX MkII imaging cytometer was performed in the Columbia University Stem Cell Initiative Flow Cytometry core facility at Columbia University Irving Medical Center and was supported by the Office Of The Director, National Institutes Of Health under Award Number S10OD026845. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.