Enzyme Chiller and Magnetic Separator for the Automated 454 Zymark/Caliper Protocol

 

Doug White and Graham Wiley

Roe Lab, University of Oklahoma

September 1, 2008

 

Enzyme chiller

 

The enzyme chiller was assembled with silicone rubber from a red anodized aluminum 96 well plate holder, a black CPU heatsink (screwed to the bottom of the auluminum holder), a tip box lid, a microtiter plate lid, and some Swagelok fittings.

 

One tube goes into 6L of ice water in an ice chest. The other tube goes to a small submersible pump in the same ice chest. The pump pulls cold water through the chiller. If a leak develops, air will be pulled into the chiller and the SciClone workspace will stay dry.  

 

 

Magnetic separator

 

The magnetic separator was assembled with silicone rubber from 117 (9x13) 0.125"x0.75" NdFeB magnets from K&J Magnetics, two 96 well viper plates, four 1/4" screws, and a microtiter plate lid.

 

The two viper plates were cut to form a 9x13 grid. Silicone was squirted into each well and one magnet was inserted in each well. A flat aluminum plate was placed on the magnets while the silicone set up. The upside down 9x13 grid was then attached to the microtiter plate lid with silicone rubber.

 

Alternating the polarities of the magnets gives a better separation of the magnetic beads and helps the magnets stay in the wells while the silicone sets up. When a 96 well (8x12) reaction plate is placed on the magnetic separator each well is surrounded by four magnets.

 

Wells A4, A10, B1, B13, H1, H13, J4, and J10 of the magnetic separator were cut off flush with the magnets to make room for the reinforcing webs on the bottom of the 96 well v-bottom reaction plates used during the prep.

 

Four holes were drilled through the magnetic separator where wells B1, B12, G1, and G12 of the reaction plate would be located. The four 1/4" plate height adjustment screws were screwed up through the bottom of the magnetic separator. A reaction plate was placed on the magnetic separator. The screws were adjusted so that the reaction plate was level and the bottoms of wells B1, B12, G1, and G12 of the reaction plate rested on top of the screws.