使用一滴附件对λDNA进行定量分析

对于许多生物样本来说，使用微量体积进行结构和定量研究更方便、更经济、更高效。SAF-850 One Drop是一种微取样附件，仅用5µL的样品就能获得荧光测量。高度可重复性的基线允许进行简单的光谱测量或对多个样品的定量分析。结论本应用说明通过获得在大浓度范围内具有良好线性的校准曲线，证明了一滴微采样附件的测量重现性。Application NoteFP-0010 Quantitative Analysis of 入 DNAUsing a One Drop Accessory2/3Application Note Quantitative Analysis of a 入 DNA Using a One Drop Accessory Introdu c tio n For many biological samples， it is much more conve n ient ， cost effective， and ef f i c i ent to use microvolumes fo r structural a n d qua n ti t ative stu d ies.The SAF-850 One Drop is a microsampl i ng accessory tha t enables fluoresce n ce measu r ements to be obtained with only 5 p L of s a mple.The h ig h ly reproduc i ble base li ne allows for either s i mple spectrum measurements or qua n t i tative analysis of mu l t ip le samples . This application note demonstrates the use of the One Drop accessory to obtain fluorescence data a n d create a calibration curve of 入DNA labeled with PicoGreen@. FP-8200 S p ec tr ofl u o r o m ete r Keyword s FP-8200， F l uorescence， Microsampling， SAF-850 One Drop Experimen t al 28600 Ma r y 's C ou r t， Easton， M D 21601 US A Figure 1. Flow char t i l l u st r a t i n g th e sa m p l e p r e p arat i on procedure us i ng t h e One Dro p acc e ssory. R esults The emission spectra of 入DNA for a variety of concentrat i ons is shown in Figure 2 and indicates t h at the maximum emission wavelength i s 523nm. Figure 2. E mis s io n sp e ctra of 入D N A labe le d w it h P ic oGr ee n @ 28600 Mary's Cour t ， E a s to n ， M D 21601US A UASGO T h e f luor e sc e n c e inte n sity a t the m axim u m emission wav ele ngt h (523 nm) was th e n m e asur e d f i ve t imes f or e ach sample to e ns u r e measur e me n t r e producibi li ty. Ta b le 1 shows th e se mea s ur e ment reproduc i bility r e sult s f or e ac h sampl e concentrat i on. Table 1. Measureme n t r eproduc i b i l i ty r esults. Concentration[ng/mL] 1 2 3 4 5 Average SD CV(%) 53.3 52.4 55.9 53.1 55.2 54.0 1.49 2.8 1 63.6 68.1 65.9 66.5 65.1 65.8 1.68 2.6 5 110.3 106.7 105.1 104.0 110.0 107.2 2.86 2.7 10 157.6 155.5 156.1 153.0 151.7 154.8 2.39 1.5 50 447.1 465.3 460.2 455.8 469.0 459.5 8.56 1.9 100 865.9 856.9 848.3 850.6 853.9 855.1 6.86 0.8 500 3842.7 3831.0 3858.0 3828.9 3811.0 3834.3 17.42 0.5 1000 7766.2 7992.1 7925.3 7972.8 7805.7 7892.4 101.15 1.3 A cal i bra t io n curve was the n created using the average value for each conce n t r at i o n from the reproducibility results in Ta b le 1 and i s s h own i n Figure 3. T h e e q u at i on of t h e lin e f it t o t he calibration curve i s y=7.780·c +57.5211. The correlat i on co e fficie n t is 0.9999 a n d th e sta n dard e rror is 6.819， ind i ca t in g a good f it . Figure 3. C al i b r at i o n curv e o f 入DNA lab ele d w ith P icoGr een@ Conclusion This a pplication n ote demonst r ates t he measuremen t r eproducibility of the One Drop microsam p ling accessory by obtaining a cal i brat i on curve with good linea r i ty over a wide concentration range. 28600 Mary's Cour t ， E a s to n ， M D 21601US A