MicroStation System, MicroLog Version 4.2 - DTU Systems Biology ...

Preparing Samples
aids in liquefying the cell mucous. Swirl the contents of the
tube occasionally during the incubation. Look for the
disappearance of the distinct mucoid strings as evidence that
the sample is ready.
Settling of the debris is critical to eliminating clumps
and obtaining a homogeneous cell suspension for
accurate identification. This step may take up to 20
minutes.
Section 4 � Page 12 MicroStation System/MicroLog User Guide Apr 07
OR
Method 2: Filter the cell mixture through a 70 µm cell
strainer (BD Falcon 352350) to pull out the cell debris
from the homogeneous cell suspension.
g. Prepare a homogeneous cell suspension. Place a new tube of
GN/GP-IF into the Biolog Turbidimeter. Adjust the zero of the
turbidimeter for the tube. Carefully remove the homogenous cell
suspension using a transfer pipette (Biolog P/N 3019) making sure
that the cell debris in the bottom of the tube is not taken. Add the
cell suspension drop wise, with mixing to the tube of GN/GP-IF in
the turbidimeter. Use a swab to stir and mix the cell suspension
into the GN/GP IF. Continue to add cells to achieve a cell turbidity
of 28% ± 2%. Uniformly suspended cells typically give stable
turbidimeter readings whereas remaining clumps and strings cause
the turbidimeter to fluctuate as they pass across the light path.
Remaining clumps can be removed by letting the tube stand until
the clumps settle. Remove the uniformly suspended cells to
inoculate the MicroPlate.
h. If more cells are needed to get the required cell density, repeat the
dry tube method preparation using a new empty, sterile dry glass
tube. Do not use the same tube that is wet with GN/GP-IF.
6. Inoculate a GP2 MicroPlate following the procedure in the Instructions
for Use.
7. Incubate at 30°C for 4 – 6 hours and/or 16 – 24 hours to allow the pattern
to form.