Supernatant Collection System

 
Cytotoxic T Lymphocytes (CTL) kill target cells on the basis of cell surface antigen recognition, and are important in the host's response for tumors, transplants, and viruses. CTLs react with a wide variety of antigens, including minor and major histocompatibility alloantigens and antigens expressed by synergetic cells, such as from haptens, tumor-associated products, and viruses.

The Supernatant Collection system is designed for collecting 90% of the supernatant in a microplate well and separating the living cell with no stress, eliminating centrifugation and other similar techniques. It can be used in a variety of release assays with different radioactive isotopes, such as Cr⁵¹ or I¹²⁵.

The Supernatant Collection system offers a safe way to handle samples containing isotopes, and is being used in cytotoxicity testing for measurement of toxic effects on cellular material and chromium release studies. By eliminating tedious centrifuging and pipetting steps in these assays, the Supernatant Collection system offers a faster, safer, and more reproducible technique, producing more reliable results. In 1978, the Supernatant Collection system was added to the National Institute of Health's (NIH) Manual of Tissue Typing Tests.

The Supernatant Collection System consists of:

1. Harvesting Frame (9000-0308)—cellulose acetate absorption cartridges with a glass fiber filter disc on the bottom of each cartridge.
2. Macrowell Tube Strips (9000-0309)—1 ml polystyrene tubes in strips of twelve that fitinto the Macrowell holder.
3. Macrowell Holder (9000-0306)—eight macrowell tube strips fit into the holder to make a 1 ml deep well plate.
4. Harvesting Press (9000-0305)—pushes the harvesting frame into the well of the round-bottom microplate containing the cell culture to be measured.
5. Transfer Fork (9000-0307)—transfers the harvesting frames containing the absorbed supernatant into the Macrowell tubes.

how the system works

The harvesting frame contains 48 cellulose acetate absorption cartridges positioned above 48 glass fiber filter discs. A frame fits over one half of a microplate so that each disc and cartridge is centered over each well (Figure 1).

		Figure 1. Discs and cartridges centered over microplate wells. 1 Absorption cartridge 2 Frame 3 Filter disc 4 Microplate well 5 Supernatant 6 Cells

The harvesting press pushes the cartridges and discs to a uniform depth in the microplate wells. The filter discs are slightly larger in diameter than the microplate wells, and thus form a seal, which traps intact cells beneath, while up to 90% of the supernatant passes through into the absorbing cartridge (Figure 2).

		Figure 2. Removing the supernatant. 1 Absorption cartridge 3 Filter disc 6 Cells 7 Press

The harvesting frame, with the absorption cartridges containing the supernatant, is lifted from the microplate. The glass fiber filter discs, with the cells trapped beneath, remain in the wells (Figure 3).

		Figure 3. After the harvesting frame is removed, the cells remain in the wells. 1 Absorption cartridge 3 Filter disc 6 Cells

The lower ends of the absorption cartridges are inserted into Macrowell™ tube strips aligned in a matching configuration in the Macrowell tube strip holder. The cartridges are then pushed to the bottom of the tubes, four at a time, using the transfer fork (Figure 4). Macrowell tubes snap apart and may be placed directly into a gamma counter, or inserted into a larger tube for counting, depending upon the counter.

		Figure 4. Depositing the cartridges into Macrowell tubes. 8 Transfer fork 9 Macrowell tube 10 Macrowell tube strip holder