Bio-Rad Bio-Dot® and Bio-Dot SF Microfiltration Apparatus - Manual (Page 8)

5. Place the sample template on top of the membrane. The guide pins ensure that the template
will be properly aligned. Finger-tighten the four screws. When tightening the screws, use a
diagonal crossing pattern to ensure uniform application of pressure on the membrane surface
(see Figure 2).
Fig. 2. Diagonal crossing pattern for tightening screws in the Bio-Dot apparatus.
6. Attach a vacuum source (house vacuum or vacuum pump) to the flow valve with a waste trap
set up and positioned between the vacuum outlet and flow valve. Turn on the vacuum and
set the 3-way valve to apply vacuum to the apparatus (flow valve setting one, Figure 3).
7. With vacuum applied, repeat the tightening process using the diagonal crossing pattern.
Tightening while vacuum is applied ensures a tight seal, preventing cross contamination
between slots. Failure to tighten screws during application of vacuum prior to starting
the assay may lead to leaking between the wells.
8. Adjust the flow valve so that the vacuum manifold is open to air (flow valve setting two,
Figure 3). Apply 100 µl to all the sample wells. Use of an 8-channel pipet and buffer
reservoirs (see Section 13 for information) will simplify the process of adding solutions to the
Bio-Dot SF apparatus. Addition of buffer is necessary to rehydrate the membrane following
the vacuum procedure in step 7. If this step is not performed prior to applying samples, assay
results will show halos or weak detection signal.
9. Gently remove the buffer from the wells by vacuum (flow valve setting three, Figure 3).
Watch the sample wells. As soon as the buffer solution drains from all the wells, adjust the
flow valve so that the unit is exposed to air and disconnect the vacuum. At this point, the unit
is ready for sample application.
Flow Valve Setting 1.
The vacuum manifold is exposed to the vacuum
source only. Use for applying vacuum to the
Bio-Dot SF apparatus.
Flow Valve Setting 2.
The manifold is exposed to air.
Use for gravity filtration procedures.
Flow Valve Setting 3.
The manifold is exposed to both air and the
vacuum. Use this setting for gentle vacuum
applications where the amount of vacuum
is regulated by putting a finger over the port
exposed to the atmosphere.
Fig. 3. Optional settings for the 3-way flow valve to obtain optimal performance from the Bio-Dot SF apparatus.
4
Vacuum Bio-Dot
Air
Air
Vacuum
Vacuum
Bio-Dot
Bio-Dot
Air