BD FACSAria II cell sorter

Frequently Asked Questions: Optics

No. The lasers in the BD FACSAria II do not require special cooling or power. The unique optical design of the BD FACSAria II allows for the use of lower-powered, air cooled, and solid state lasers for efficiency, without sacrificing sensitivity or resolution.

The excitation optics in the BD FACSAria II consist of lasers, fiber optic cables, beam-shaping prisms, and an achromatic focusing lens. The fiber optics direct the laser light from the 488-nm, 633-nm, and 405-nm lasers precisely into beam-shaping prisms that transmit the laser light into a focusing lens. The lens then focuses the laser light into the cuvette flow cell. The 375-nm laser is air launched and focused onto the cuvette flow cell.

No. Since the optical pathway and the sample core stream are fixed, alignment is constant from day to day.

The collection optics are set up in octagon- and trigon-shaped arrays that maximize signal detection from each laser. This is accomplished by transmitting the longest wavelengths to the first photomultiplier tube (PMT), and reflecting shorter wavelengths to each subsequent PMT through a series of longpass dichroic mirrors. Bandpass filters in front of each PMT allow fine-tuning of the spectral wavelengths that need to be collected.

The design is based on the fact that light reflection is more efficient than light transmission. Emitted light travels to each PMT detector via reflection and is transmitted through only two pieces of glass to reach the detectors. This greatly increases sensitivity and resolution for each color in a multicolor panel.

Having the cuvette gel-coupled to the fluorescence objective lens allows for the greatest amount of emitted light to be transmitted from the interrogation point to the collection optics, and improves sensitivity and resolution.

To provide maximum flexibility, the BD FACSAria II cell sorter can be configured with up to four air cooled, solid state lasers and can detect up to 15 parameters simultaneously (13 fluorescent markers and 2 scatter parameters) when fully equipped.

A standard system is equipped with an octagon containing six PMTs that detect light from the 488-nm (blue) laser, and a trigon containing two PMTs that detect light from the 633-nm (red) laser. The third laser option adds an additional trigon containing two PMTs to detect light from the 405-nm (violet) and/or 375-nm (Near UV) lasers. Standard instruments can be configured with up to four lasers. The violet and Near UV laser share the same beam spot at the point of interception within the cuvette, so the user must select to run either the violet laser, the Near UV laser, or neither.

The 375-nm Near UV laser option has been designed to sort and analyze mouse and hematopoietic stem cells.

Measuring only 19 × 7 × 7 cm, the fully-housed 375-nm Near UV laser fits in the small space immediately left of the flow cell. The laser produces a 10-mW focused beam to illuminate the cells in the flow cell.

The top hat design of the 375-nm Near UV laser modifies the usual Gaussian beam that illuminates a sample at higher intensity in the middle of the beam and lower intensity at the edges. The composite “square” beam illuminates the middle and edges of the sample core stream at the same intensity. The square beam triples the amount of light at the sample intercept.

Back to Frequently Asked Questions »

BD flow cytometers are Class I (1) laser products.

For research use only. Not for use in diagnostic or therapeutic procedures.