Laser Enabled Analysis and Processing (LEAP) is a system that combines high-speed whole well imaging and ultra fast laser manipulation for selective and efficient processing of cells in their natural state. These capabilities enable cell purification, enrichment, cloning, stem cell colony processing and optoinjecection (transient permiabilization of cells).

LEAP has been used for a variety of applications including:

• Purification of human embryonic (ES) and induced pluripotent (iPS) stem cells
• Purification of cancer stem cells
• Purification of primary cells with both high yield and high purity, (e.g. brain microvascular endothelial cells)
• Rapid stable cell line development in 3 weeks (e.g. GPCR expression, reporter genes)
• Rapid stable secretion cell line development
• Delivery of a variety of molecules into cells (e.g. siRNA, peptides/proteins, quantum dots, molecular biosensors, etc.)

LEAP is an in situ cell purification system with the following benefits:

• Clone or enrich populations based on cell phenotype
• Purify adherent or non-adherent cells
• Consistent high yield and high purity
• Purify small samples (10’s -> 10⁶)
• Purify directly within multi-well plates
• Closed system suitable for biohazardous samples
• Image-based validation of purification
• Process cells directly in growth medium
• Process cells in sterile, closed plate environment
• Easily automatale high throughput and scalablity

For cell purification, viability of the purified cells is not affected since the laser does not touch them. Only the health of the cells impacted by the laser is affected. During optoinjection where the laser does directly impact the desired cells, cell yield will depend upon the cell type and laser conditions. Typically, cell yields vary from 50 – 90% during optoinjection under optimal conditions.

We would be happy to present results from practical implementations of LEAP-based cell manipulations.

LEAP uses SBS-standard format multi-well plates including 384, 96, 12, and 6 well plates.

Cyntellect has designed specific multi-well plates with specific tolerances to be used with LEAP. C-lect™ plates include different well densities that are compatible with the SBS standard footprint. These plates will withstand laser manipulation with minimal damage compared to conventional plates and are optimized for current applications. We continue to expand our plate offerings.

The LEAP system includes:

• Whole well imaging (96W and 384W plates)
• Brightfield illumination (LED-based)
• Fluorescence imaging (8 position filter wheels)
• Hybrid-halide lamp excitation
• Mega-pixel emCCD camera
• Adjustable magnification (3, 5, 10, 20×)
• Rapid auto-focus
• Real-time gating interface
• Laser-based cell manipulation (at 355 and 532 nm)
• Cell targeting at >1,000 per second

The following Label-Free Assays for Live Cells can be used on the Celigo cytometer

• Cell counting for adherent cells and for non-adherent cells
• Cell growth tracking
• Colony counting and confluence measurement

The following fluorescence assays can be used on the Celigo cytometer

• Cell counting
• Cell viability (toxicity)
• DNA synthesis (cell proliferation)
• PS externalization (apoptosis)
• Cell Secretion Assay (Measurement of antibody secretion from individual cells)

The Celigo cytometer has 3 excitation filters and 3 emission filters, to allow for 1, 2, or 3 channels of fluorescence data. Multiple channels of fluorescence data can be a combination of brightfield and fluorescence, which will allow 3 fluorescence and 1 brightfield channels.

The Celigo cytometer can accommodate the following types of plates • 384, 96, 24, 12, 6 well plates (1536 well plate format available soon). It can also accommodate T-25 and T-75 flasks

The data image export file format is TIFF and the image format for database storage is JPEG 2000. Data can be exported as .csv files and excel files also.