( ove cell growth. 3T3 cells (ATCC Rockv...)Direct Counting of Millipore MultiScreen Filtration Plates.

ove cell growth. 3T3 cells (ATCC, Rockville, Maryland) were cultured to 104 cells per well and stimulated with Bovine Pituitary Extract (Collaborative Biomedical Products, #40123) at levels ranging from 0 to 250 ug/mL for 18 hours. The cells were then pulsed with ³H-thymidine for three to four hours prior to washing on the MultiScreen vacuum manifold. Two plates were prepared. The first was processed by punching the individual membrane disks into LS vials, solubilizing with 0.42% bleach, and mixing with 5 mL LS cocktail (Optifluor, Packard Instrument Company). The second plate was processed by adding 20 mL MicroScint-O to each well using a semi-automatic multichannel pipet and counting on TopCount.

Stimulation curves obtained with the TopCount and with conventional LSC are shown in Figure 4. Both methods yield the same results.

Other Assays.
A variety of other assays such as enzyme inhibition and kinase assays have been performed with the MultiScreen filtration plate on TopCount. Counting efficiencies for the radionuclides used in these assays are summarized in Table 2. For weak beta emitters such as ³H, ³⁵S, and ¹⁴C, crosstalk was negligible. For gamma emitters (¹²⁵I) and higher energy beta emitters (³²P), crosstalk was less than 0.5%.

Conclusions

The Millipore MultiScreen Assay system is designed for a variety of microplate assays requiring in plate incubations and filtration. A significant limitation of the system is the need to individually process the 96 samples in a plate for quantitation. The TopCount Microplate Scintillation Counter can facilitate the processing of MultiScreen filtration plates by allowing the user to process all 96 samples as a single unit. Crosstalk due to physical migration of radiolabel is not possible due to the discrete filter disks i
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