研究目的
To investigate the molecular impact of antimicrobial blue light (aBL) treatment on methicillin-resistant Staphylococcus aureus (MRSA), focusing on changes in intracellular porphyrin, reactive oxygen species, and fatty acid profiles to uncover the bactericidal mechanism.
研究成果
aBL irradiation causes excitation of intracellular coproporphyrin, leading to generation of singlet oxygen and ROS, which oxidize unsaturated fatty acids in the cell membrane, resulting in membrane damage, increased permeability, and bacterial death. This provides quantitative evidence supporting the hypothesis that the cell membrane is a key target in aBL's bactericidal mechanism.
研究不足
The study is limited to in vitro conditions using a specific strain (MRSA252) and may not fully represent in vivo scenarios or other bacterial strains. The mechanisms identified are correlative and require further validation, such as through lipidomic studies or genetic manipulations.
1：Experimental Design and Method Selection:
The study used a 415-nm LED light source to irradiate MRSA252 cells at various doses (0-240 J/cm2) to assess bactericidal effects and sub-lethal changes. Methods included HPLC for coproporphyrin, CLSM for singlet oxygen, spectrofluorophotometry for ROS, colorimetry for MDA, GC-MS for fatty acids, AFM for morphology, and flame atomic absorption spectrometry for K+ leakage.
2：Sample Selection and Data Sources:
MRSA252 strain was grown on LB agar and in broth, with cell suspensions prepared in PBS at OD600 of 0.5 for experiments.
3：5 for experiments. List of Experimental Equipment and Materials:
3. List of Experimental Equipment and Materials: Equipment included a 415-nm LED array (Omnilux clear-U), power meter (PM100D), centrifuge, HPLC system, CLSM (TCS SP8, Leica), spectrofluorophotometer, AFM, and flame atomic absorption spectrometer. Materials included SOSG reagent, DCFH-DA probe, MDA assay kit (Beyotime #S0103), ROS assay kit (Beyotime #S0033), and various chemicals like PBS, NH4OH-acetone solution, and BSTFA.
4：Experimental Procedures and Operational Workflow:
Cells were irradiated in a 6-well plate with stirring, sampled at specific time points, and analyzed for survival, coproporphyrin, ROS, MDA, fatty acids, morphology, and K+ leakage using standardized protocols with controls and statistical analysis (ANOVA).
5：Data Analysis Methods:
Data were analyzed using one-way ANOVA with p<0.05 considered significant, and results expressed as mean ± standard deviation from triplicate experiments.
独家科研数据包，助您复现前沿成果，加速创新突破
获取完整内容-
Confocal laser scanning microscope
TCS SP8
Leica Microsystems Inc.
Used for imaging singlet oxygen with SOSG reagent.
-
LED array
Omnilux clear-U
Photo Therapeutics, Inc.
Used as the blue light source for antimicrobial irradiation.
-
Power meter
PM100D
Used to detect and measure the irradiation level.
-
ROS assay kit
S0033
Beyotime Biotech
Used to detect intracellular reactive oxygen species.
-
MDA assay kit
S0103
Beyotime
Used to quantify malondialdehyde as a marker of lipid peroxidation.
-
Atomic force microscope
Used to observe changes in cell surface morphology.
-
Flame atomic absorption spectrometer
Used to measure extracellular K+ concentration.
-
GC-MS
Used for fatty acid profiling.
-
HPLC system
Used to detect coproporphyrin levels.
-
Spectrofluorophotometer
Used to measure ROS levels with DCFH-DA probe.
-
登录查看剩余8件设备及参数对照表
查看全部
