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Amnis ImageStream MkII

The Amnis ImageStreamX MkII is a benchtop imaging flow cytometer system that combines the detailed cellular images and morphologic information of microscopy with the large sample size and phenotyping abilities of flow cytometry. The system has two CCD cameras, three lasers (Violet 405 nm, Blue 488 nm, Red 642 nm), and three imaging objectives (20x, 40x, 60x). The ImageStreamX MkII captures multiple images of each cell in flow, including brightfield, darkfield, and up to 10 fluorescent markers simultaneously. The image resolution depends on the magnification settings, with pixel sizes of 1 μm2, 0.25 μm2, and 0.1 μm2, respectively.

Data analysis is performed using the IDEAS software, which utilizes wizard-based and customized workflows to objectively quantify distinct cell populations, cellular features, and processes, including nuclear translocation, shape change, internalization, and apoptosis.

Applications include:

  • Intracellular co-localization
  • Nuclear translocation
  • Internalization and phagocytosis
  • Cell-cell interaction
  • Cell death and autophagy
  • Cell cycle and mitosis

For more information, visit the manufacturer's website: https://cytekbio.com/pages/imagestream

When planning an imaging flow cytometry experiment and choosing fluorochromes, it is important to know the fluorochromes' specific excitation and emission wavelengths, as well as the lasers, filters, and detectors available on the instrument. Below is the information for the Amnis ImageStreamX MkII.

Amnis ImageStreamX MkII Configuration and Fluorochromes
CameraExcitation LaserChannelBandpassFilterIdeal dyesAdditional Fluorochromes
Camera 1Bright fieldCh1 Bright FieldBright Field
Blue 488 nm  Ch2528/65 (480-560)FITC, AF488, GFPCFSE, YFP, DyLight488, PKH67-membrane label, LysoTrackerGreen
Ch3577/35 560-595PECy3, DSRed, PKH67-membrane label SYTOX Orange
Ch4610/30 595-642PE-TexasRed*RFP, PE-AF610, eFluor625, Qdot625* Ch4 & Ch10 BF**
Ch5702/85 642-745PE-Cy5*, PE-AF647PI, 7AAD, PerCP-Cy5.5, eFluor 650, Draq 5
Ch6765/40 745-785PE-Cy7*, PE-AF750or use for side scatter (SSC)
Camera 2  Violet 405 nmCh7457/45 435-505DAPI, AF405, BV421Hoechst, eFluor 450, DyLight 405, CFP, Live/Dead violet
Ch8537/65 505-570BV510AF430, eFluor506, Qdot 525, Cascade yellow
Ch9582/25 570-595Bright Field Qdot565, Qdot585, eFluor565
Ch10610/30 595-642 Qdot 605, Qdot625, eFluor625, BV605 Ch4 & Ch10 BF**
Ch11702/85 642-745 BV711, eFluor700 Qdot 705*
Ch12765/40 745-785 Qdot 800*, BV786 or use for side scatter
Red 640 nmCh11702/85 642-745AF647, AF660, AF680APC, Cy5, AF660, Draq 5*, DyLight649, DyLight680, PE-AF647*, PE-Cy5
Ch12765/40 745-785APC-Cy7, APC-AF750APC-H7, Cy7, APC-eFluor780, AF-750, DyLight750, PE-Cy7* PE- AF750* or use for side scatter

*It is important to remember that many fluorescent dyes are excited by more than one laser and fluoresce in multiple channels. It is very helpful to use a spectral viewer when planning an experiment.

** There are options to switch the Brightfield to different channels if needed (Ch1 & Ch9, Ch2 & Ch8, Ch 4 & Ch10, and Ch6 & Ch12).

Ideal fluorochromes for this instrument are AF488, PE, PE-TexasRed, SSC-Ch6, DAPI (or BV421), AF647, and APC-Cy7.