Validation of automated spectrofluroimetry for measurement of regional organ perfusion using fluorescent microspheres.
Validation of automated spectrofluroimetry for measurement of regional organ perfusion using fluorescent microspheres.
复制标题
使用荧光微球测量区域器官灌注的自动荧光光谱法的验证。
DOI：
10.1016/s0169-2607(00)00057-2
复制
发表时间：
2000
影响因子：
6.1
通讯作者：
Glenny,RW
中科院分区：
文献类型：
--
作者：
Schimmel,C;Frazer,D;Huckins,SR;Glenny,RW
用于测定局部器官血流的荧光微球（FM）方法是劳动密集型的，需要处理每个器官100 s至1000 s的样品。为了节省时间和减少操作错误，我们开发了一种自动荧光分析系统，将市售荧光分光光度计和样品输送单元连接到PC上，由基于Windows95®的程序WINFAC操作。WINFAC允许多功能的分析设置和仪器控制，用于在固定波长或通过同步扫描进行荧光强度采集。数据实时显示在屏幕上，并以文本格式存储。在规定的时间间隔进行参比分析，并不断更新变异系数以监测仪器性能。针对放射性微球（RM）在猪和羊肺灌注的自动化系统进行了验证，并评价了分析重现性。通过同时注射到股静脉中递送荧光和放射性微球。切除肺，冲洗，干燥至总肺容量，并切成1.2cm 3的小块（n=833和1560，分别为猪和羊）。测定每个肺片的放射性计数率（校正衰变、背景和溢出）。在溶剂中提取荧光染料，并使用自动荧光分光光度计（校正背景和溢出）在固定波长下测定强度。每50个样品测量多色参比溶液，以监测仪器再现性。通过RM和FM方法确定的每件血液流量估计值高度相关：R2=0.98±0.017，斜率=1.00±0.007，截距=0.00±0.006（平均值±SD）。重复参考分析的CV为0.71%±0.16，相对于手动分析降低了30%至50%。自动荧光分光光度法减少了测量误差，是一种可靠和节省时间的进步。利用该技术，使用FM测量局部肺灌注接近RM方法的简便性和准确性。
The fluorescent microsphere (FM) method for determination of regional organ blood flow is labor intensive, requiring processing of 100 s to 1000 s of samples per organ. To save time and reduce handling errors, we developed an automated fluorescence analysis system by interfacing a commercially available spectrofluorimeter and sample delivery unit to a PC, operated by a Windows95®-based program, WINFAC. WINFAC allows versatile analysis setup and instrument control for fluorescent intensity acquisition at fixed wavelengths or by synchronous scanning. Data is presented on screen, in real-time, and stored in text format. Reference analyses are performed at specified intervals and the coefficient of variation is continuously updated to monitor instrument performance. The automated system was validated against radioactive microspheres (RM) for lung perfusion in a pig and sheep and evaluated for analysis reproducibility. Fluorescent and radioactive microspheres were delivered by simultaneous injection into a femoral vein. Lungs were excised, flushed, dried at total lung capacity, and cubed into ≈2cm3pieces (n=833 and 1560, pig and sheep, respectively). Radioactive count rates were determined for each lung piece (corrected for decay, background and spillover). Fluorescent dyes were extracted in solvent and intensities were determined at fixed wavelengths, using the automated spectrofluorimeter (corrected for background and spillover). Multi-color reference solutions were measured every 50 samples to monitor instrument reproducibility. Blood flow estimates for each piece determined by RM and FM methods were highly correlated: R2=0.98±0.017, Slopes=1.00±0.007 and Intercepts=0.00±0.006 (mean±SD). The CV of repeat reference analyses was 0.71%±0.16, a 30% to 50% reduction relative to manual analysis. Automated spectrofluorimetry reduces measurement errors and is a reliable and time saving advancement. With this technology, use of FM to measure regional lung perfusion approaches the ease and accuracy of the RM method.