NaOH-SDS溶液结合流式细胞术快速鉴别革兰阳性、阴性菌

niwanmao

用NaOH-SDS溶液作用细菌溶液5分钟，几乎完全溶解的是大肠杆菌等革兰阴性菌，而很难溶解的是粪肠球菌等革兰阳性菌。
使用的仪器是尿颗粒分析仪UF-1000i（原理与流式细胞仪类似）

NaOH-SDS溶液作用于不同细菌

PLoS One. 2012;7(10):e47093. doi: 10.1371/journal.pone.0047093. Epub 2012 Oct 15.
Rapid Discrimination of Gram-Positive and Gram-Negative Bacteria in Liquid Samples by Using NaOH-Sodium Dodecyl Sulfate Solution and Flow Cytometry.
Wada A   , Kono M   , Kawauchi S   , Takagi Y   , Morikawa T   , Funakoshi K   .
Source
Cell Analysis Center, Scientific Affairs, Sysmex Corporation, Nishi-ku, Kobe, Hyogo, Japan.

Abstract
BACKGROUND:
For precise diagnosis of urinary tract infections (UTI), and selection of the appropriate prescriptions for their treatment, we explored a simple and rapid method of discriminating gram-positive and gram-negative bacteria in liquid samples.
METHODOLOGY/PRINCIPAL FINDINGS:
We employed the NaOH-sodium dodecyl sulfate (SDS) solution conventionally used for plasmid extraction from Escherichia coli and the automated urine particle analyzer UF-1000i (Sysmex Corporation) for our novel method. The NaOH-SDS solution was used to determine differences in the cell wall structures between gram-positive and gram-negative bacteria, since the tolerance to such chemicals reflects the thickness and structural differences of bacterial cell walls. The UF-1000i instrument was used as a quantitative bacterial counter. We found that gram-negative bacteria, including E. coli, in liquid culture could easily be lysed by direct addition of equal volumes of NaOH-SDS solution. In contrast, Enterococcus faecalis, which is a gram-positive bacterium, could not be completely lysed by the solution. We then optimized the reaction time of the NaOH-SDS treatment at room temperature by using 3 gram-positive and 4 gram-negative bacterial strains and determined that the optimum reaction time was 5 min. Finally, in order to evaluate the generalizability of this method, we treated 8 gram-positive strains and 8 gram-negative strains, or 4 gram-positive and 4 gram-negative strains incubated in voluntary urine from healthy volunteers in the same way and demonstrated that all the gram-positive bacteria were discriminated quantitatively from gram negative bacteria using this method.
CONCLUSIONS/SIGNIFICANCE:
Using our new method, we could easily discriminate gram-positive and gram-negative bacteria in liquid culture media within 10 min. This simple and rapid method may be useful for determining the treatment course of patients with UTIs, especially for those without a prior history of UTIs. The method may be easily applied in order to obtain additional information for clinical prescriptions from bacteriuria.
PMID: 23077549 [PubMed - in process] PMCID: PMC3471971

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