Biomedical Engineering Reference
In-Depth Information
PEG groups. Consider three diferent ways of producing the cellular micropattern: (a) using
photolithography, (b) using microcontact printing, and (c) using microluidic patterning. In all
cases, the techniques must be used to deine the ECM and PEG micropatterns (not to directly
deposit the cells). Explain the biological basis of why the cells prefer to attach to the ECM protein
micropattern and never spread beyond, into the PEG areas.
Exercise A.2.6 (Design Challenge).
“A microfabricated clonogenic assay.” he goal of this
experiment is to measure the replication rates of approximately 100,000 single cells. You are part
of a large research team, but your job is the most critical one. You are asked to design a microar-
ray of single cells on a substrate that will allow you to visualize the cells dividing in real time
using a microscope. Optimization of the time-lapse microscope is not your job, so you work
under the assumption that the cells, under the microscope, will be living and dividing happily.
Another person in your research team will take care of counting the cells from the images you
acquire, so you don't need to worry about that either. Your job is only to design a substrate on
which you will be seeding a suspension of adherent cells that a technician will hand to you. (You
can choose a cell type if you want.)
Remember that as the cells divide, the “colony” will grow in size—ater the irst division,
you won't have single cells, you will have pairs of single cells, and so on. You are only going
to acquire data for the irst 10 divisions. You have one design constraint: the colonies can-
not overlap for the duration (10 divisions) of the experiment. You inally decide that you
will try two diferent microfabrication techniques (no robotics to position anything, please).
Describe them in detail, drawing the cross-section schematics for clarity if you think you
need to.
Obviously, you have to make a combination of design choices on the type of substrate,
substrate coatings, and microfabrication technique. Challenge yourself by choosing two
combinations that work
and
correctly explaining why you made those choices. Instead of
using vague, obvious statements such as “we will use a cell-adhesive protein,” specify which
cell-adhesive protein. Similarly, do not say “albumin will be immobilized”—instead, specify
the physical principles or chemical reactions (or both) you used to immobilize it, and instead
of indicating “we will micropattern collagen,” specify which micropatterning technique you
used.
Exercise A.2.7.
To promote immobilization of proteins on a Si-OH modiied surface, you
would like to use an epoxy functional silane coating. You have your choice of a (3-glycidyloxy-
propyl)trimethoxysilane and (3-glycidoxypropyl)methyldiethoxysilane. How many reactive
groups does each silane have for proteins? For the surface? For other silanes? Which do you
think will be most efective at making a stable and cross-linked surface coating?
Exercise A.2.8.
Approximate the following molecular species of interest for surface coat-
ing by simple geometric shapes and calculate the maximum packing density (molecules/cm
2
),
assuming a minimum intermolecular spacing of 0.5 Å
(a) Alkanethiol—rod, 10 Å length, 1 Å diameter
(b) Bovine serum albumen (BSA)—ellipsoid, 140 × 40 × 40 Å
3
You are coating the surfaces of a closed microwell that is 100 × 100 × 50 μm
3
with the species
above. he inlet and outlet dimensions of the chamber are 50 × 50 μm
2
. he coating protocol is
to load a solution of the desired species diluted with bufer into the chamber, allow it to sit for
1 hour to coat, and then lush the chamber with bufer. Assume that the molecules will pack at
the density from part 1 of the problem, only one monolayer will form on the surface, and the
rest of the molecules will stay in solution. he concentration of molecule in the solution should
not change by more than 1% during the coating process. Calculate the minimum concentrations
necessary.
Exercise A.2.9.
A femtosecond titanium-sapphire laser, with a pulse frequency of 76 MHz and
pulse duration of 150 fs, tuned to 740 nm was used for fabrication of albumin microstructures.
