Biomedical Engineering Reference
In-Depth Information
dense biofilms. In (C), NACP
QADM had less biofilms, where “R” indicates resin composite not
covered by biofilms. Higher magnification (D, E) revealed that the S. mutans grew in chains
(arrows). The chains twisted in three-dimensions and were long or continuous in the biofilm archi-
tecture. The chains were much shorter on NACP
QADM in (F), with each chain containing 3
10
cells.
For MTT assay, disks were placed in 24-well plates, inoculated with 1.5 mL inoculation
medium, and cultured for 3 days. Each disk was transferred to new 24-well plates for the MTT
(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay
[20,21]
. MTT is a colorimet-
ric assay that measures the enzymatic reduction of MTT, a yellow tetrazole, to formazan. One mil-
liliter of MTT was added to each well and incubated for 1 h. Disks were transferred to new 24-well
plates, and 1 mL of dimethyl sulfoxide (DMSO) was added to solubilize the formazan crystals. The
DMSO solution from each well was used and the absorbance at 540 nm was measured via a micro-
plate reader (SpectraMax M5, Molecular Devices, Sunnyvale, CA).
For lactic acid measurement, disks with 3-days biofilms were placed in new 24-well plates, and
1.5 mL of buffered-peptone water (BPW) supplemented with 0.2% sucrose was added. They were
incubated for 3 h to allow the biofilms to produce acid. Then the BPW solutions were stored for
lactate analysis. The microplate reader was used to measure the absorbance at 340 nm, and standard
curves were prepared using a lactic acid standard (Supelco, Bellefonte, PA)
[21]
.
Figure 6.6
plots the results for MTT metabolic activity, CFU, and lactic acid production of bio-
films. For each property, the values did not vary significantly over aging time (P
.
0.1). In (A),
NACP
QADM
greatly reduced the CFU (B) and lactic acid (C), compared to commercial composites (P
,
QADM yielded much lower MTT than commercial composites (P
,
0.05). NACP
0.05).
Therefore, the novel NACP
QADM nanocomposite substantially decreased the biofilm growth and
lactic acid production, and its antibacterial properties were maintained during long-term water-
aging
[52]
.
6.4
Antibacterial dentin primer
Scotchbond multi-purpose (referred as “SBMP”) (3M, St. Paul, MN) was used as the parent bond-
ing agent to test the effect of QDMA and NAg incorporation. The purpose was to develop a model
system, and the novel method of QADM and NAg incorporation can then be applied to other bond-
ing agents. SBMP etchant contained 35% phosphoric acid. SBMP primer contained 35
45%
2-hydroxyethylmethacrylate (HEMA), 10
20% copolymer of acrylic/itaconic acids, and 40
50%
water. SBMP adhesive contained 60
40% HEMA. QADM and NAg were
incorporated into SBMP primer. SBMP etchant and adhesive were not modified.
Bis(2-methacryloyloxyethyl) dimethylammonium bromide (QADM) was recently synthesized
[20,21]
. Briefly, 10 mmol of 2-(N,N-dimethylamino)ethyl methacrylate (Sigma-Aldrich) and
10 mmol of 2-BEMA (Monomer-Polymer Labs, Trevose, PA) were combined in ethanol and stirred
at 60
C for 24 h. The solvent was then evaporated, yielding the QADM. QADM was mixed with
the SBMP primer at QADM/(primer
70% BisGMA and 30
10 wt%. This was selected because preliminary
study showed that 10% of QADM in the primer provided antibacterial activity without compromis-
ing the dentin bond strength, while 20% QADM in primer decreased the bond strength.
QADM)
1
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