使用带有一滴附件的紫外可见分光光度计测量再现性

SAH-769 One Drop附件用于快速、准确地定量小体积的蛋白质和核酸。为了进行测量，将单滴样品移液到细胞上（5μL为1mm或0.6μL为0.2 mm的光径长度）。Application NoteUV-O021 Measurement Reproducibility using a SAH-7692/5Application NoteOne Drop Measurement Accessory Measurement Reproducibility using a SAH-769 One Drop Measurement Accessory Introduc t ion The SAH-769 One Drop accessory is used for fast and accurate quantitat i o n smal l volumes of prote i ns and nucleic acids. To make a measurement a single drop of sample is pipetted onto the cell (5pL for 1mm or 0.6pL for 0.2 mm optical pathlengt h s). This application n ote demo n strates both the reproducibil i ty a n d l in ear i ty of the SAH-769One Drop accesso r y using cal f t h ymus DNA with both 1 and 0.2 mm pathleng th cells. V-630 UV-Vis Spectro p hotometer Keywords V-630BIO ， UV-Visible /N IR ， Bioch e mistry， SA H-769 O n e D ro p ， Mic r osampl in g Exper i m e ntal Wavelength 260nm Bandwidth 1.5nm Response Medium A dro p of sample was dis p ensed， me a su r ed ， a n d r e p eated 10 t i mes. The sample was wi p ed f rom the cell in between measureme n ts. The procedure can be see n i n Figure 1. 28600 Ma r y 's C ou r t， Easton， M D 21601 US A Figure 1. M e a su r emen t p roc e d ur e us i ng t h e S A H -769 O n e Dr op a cc e ssory . Resul t s Th e ab s o rpt io n v a lu es ob t a in ed i n a 1 mm p a th l en gth ce l l a r e sh o w n in T a bl e 1 an d the ca libr a ti o n cu r ve i n Fi g u r e 2. Ta bl e 2 i llu st r a te s t h e r e pr oducibility o f th e co n cen tr atio n val u e s ca l c u la t e d f r o m th e ab s o rb a n ce d ata. Table 1. S A H -769 a b s or b a nce m ea su r emen t rep rod u c i bil i t y u sing a1 mm p a thlengt h cell. ng/uL 0 2.4 4.8 9.6 19.3 38.6 77.2 154.4 308.8 617.5 1 -0.0008 0.0047 0.0084 0.0179 0.0321 0.0676 0.133 0.259 0.516 0.998 2 0.0003 0.0059 0.0093 0.0146 0.0332 0.0677 0.131 0.260 0.518 1.003 3 0.0012 0.0056 0.0083 0.0162 0.0334 0.0705 0.130 0.260 0.519 1.003 4 0.0015 0.0063 0.0072 0.0181 0.0345 0.0679 0.130 0.262 0.512 1.024 5 -0.0002 0.0067 0.0071 0.0166 0.0329 0.0676 0.132 0.264 0.517 0.993 6 -0.0013 0.0053 0.0088 0.0204 0.0331 0.0672 0.130 0.259 0.522 0.996 7 0.0013 0.0049 0.0082 0.0170 0.0336 0.0695 0.129 0.259 0.511 0.996 8 0.0002 0.0036 0.0089 0.0168 0.0326 0.0680 0.133 0.260 0.509 1.006 9 0.0027 0.0058 0.0089 0.0177 0.0315 0.0676 0.134 0.260 0.509 1.000 10 -0.0004 0.0043 0.0069 0.0153 0.0353 0.0692 0.132 0.267 0.509 0.995 Avg. 0.0004 0.0053 0.0082 0.0171 0.0332 0.0683 0.131 0.261 0.514 1.001 S.D. 0.0012 0.0010 0.0008 0.0016 0.0011 0.0011 0.0015 0.0026 0.0047 0.0089 C.V.% N/A 17.9 10.3 9.6 3.3 1.6 1.2 1.0 0.9 0.9 28600 Mary's Cour t ， E a s to n ， M D 21601US A Figure 2. C a l ib r at ion c ur ve c re a te d u si n g data from T a b le 1. The cal i bra ti o n equa t ion is ：Abs = 0.00163 x C onc.+0.00366 a nd t h e co r rela ti o n co e ff ic ien t is 0.9998. Table 2. C onc e nt r at i o n r e prod u ci b il i t y u si n g a 1 mm pathl e ng th cel l . ng/uL 2.4 4.8 9.6 19.3 38.6 77.2 154.4 308.8 617.5 1 -2.8 0.6 2.9 8.8 17.5 39.3 79.8 157.3 315.0 611.5 2 -2.1 1.4 3.4 6.7 18.2 39.4 78.0 157.7 316.3 614.6 3 -1.5 1.2 2.9 7.7 18.3 41.1 78.0 157.9 316.9 614.7 4 -1.3 1.7 2.2 8.9 18.9 39.5 77.7 158.8 312.4 627.6 5 -2.4 19. 2.1 8.0 18.0 39.3 78.7 160.0 316.0 608.8 6 -3.1 1.0 3.1 10.3 18.1 39.1 77.5 157.0 318.7 610.4 7 -1.4 0.7 2.8 8.2 18.4 40.5 77.4 157.1 312.0 610.3 8 -2.1 0.0 3.2 8.1 17.8 39.6 79.3 157.8 311.1 616.5 9 -0.6 1.3 3.2 8.6 17.1 39.3 80.0 157.4 310.9 612.8 10 -2.5 0.4 2.0 7.1 19.5 40.3 78.9 162.2 311.0 609.9 Avg. -2.0 1.0 2.8 8.2 18.2 39.8 78.5 158.3 314.0 613.7 S.D. 0.75 0.59 0.52 1.00 0.68 0.66 0.95 1.63 2.88 5.48 C.V.% N/A 57.4 18.7 12.2 3.7 1.7 1.2 1.0 0.9 0.9 *De t ection l imi t： 50 n g /uL 28600 Mary's Cour t ， E a s to n ， M D 21601US A The absorp ti o n va lu e s obtained in a 0.2 mm path l eng t h ce l l a r e shown in Ta bl e 3 and the calibration c u rve i n Fi g u r e 3.T a b le 4 illustr a te s the r e pr od uc ibility of the c o n c e n tr a t i o n v a lu e s c a lcul a ted from th e ab s o rb a n ce d ata. Table 3. S AH-769 absor b a n c e me a s ure men t r e p r od u cibility u sin g a 0.2 mm p a t hl e n g t h c el l . ng/uL 0 38.6 77.2 154.4 308.8 617.5 1235 2470 4940 1 0.0008 0.0122 0.0196 0.0460 0.0842 0.166 0.326 0.646 1.297 2 -0.0011 0.0165 0.0223 0.0430 0.0847 0.166 0.330 0.645 1.319 3 -0.0001 0.0160 0.0195 0.0427 0.0873 0.170 0.328 0.653 1.320 4 -0.0004 0.0112 0.0176 0.0430 0.0863 0.165 0.331 0.654 1.284 5 0.0069 0.0152 0.0180 0.0410 0.0860 0.168 0.327 0.647 1.307 6 -0.0005 0.0139 0.0184 0.0424 0.0845 0.170 0.326 0.660 1.303 7 0.0006 0.0137 0.0206 0.0450 0.0839 0.170 0.331 0.659 1.312 8 0.0019 0.0127 0.0200 0.0440 0.0875 0.168 0.330 0.658 1.301 9 0.0011 0.0130 0.0212 0.0435 0.0851 0.169 0.338 0.663 1.296 10 0.0007 0.0134 0.0194 0.0426 0.0879 0.171 0.326 0.664 1.308 Avg. 0.0010 0.0138 0.0197 0.0433 0.0857 0.168 0.329 0.655 1.305 S.D. 0.0012 0.0017 0.0015 0.0014 0.0015 0.0020 0.0037 0.0071 0.0110 CV.% N/A 12.2 7.4 3.3 1.7 1.2 1.1 1.1 0.8 T h e abso r p t i o n v a lues o b tai n ed in a 0.2 mm p a th le n g t h cel l are s h o wn i n T able 3 an d the c ali b r a ti o n cu rv e i n Fi g u re 3.Table 4 illustrates th e reprodu cib i lit y of th e conce n trat i on values calculated from th e a b sorbance d at a . Figure 3. C al i b r at i o n c urv e c re a te d us i n g data from T ab le 3. The cal i b r at i o n e quat i o n is ：A b s=0.00264x C o n c . +0.00281 an d t he c or rela t ion co e ff ic i e n t i s 0.9999. 28600 Mary's Cour t ， E a s to n ， M D 21601US A Table 4. C on c e n t r ati o n r ep ro d u c i b il it y us i ng a 0.2 mm pat h l en g th c e l l . ng/pL 0 38.6 77.2 154.4 308.8 617.5 1235 2470 4940 1 0.0008 0.0122 0.0196 0.0460 0.0842 0.166 0.326 0.646 1.297 2 -0.0011 0.0165 0.0223 0.0430 0.0847 0.166 0.330 0.645 1.319 3 -0.0001 0.0160 0.0195 0.0427 0.0873 0.170 0.328 0.653 1.320 4 -0.0004 0.0112 0.0176 0.0430 0.0863 0.165 0.331 0.654 1.284 5 0.0069 0.0152 0.0180 0.0410 0.0860 0.168 0.327 0.647 1.307 6 -0.0005 0.0139 0.0184 0.0424 0.0845 0.170 0.326 0.660 1.303 7 0.0006 0.0137 0.0206 0.0450 0.0839 0.170 0.331 0.659 1.312 8 0.0019 0.0127 0.0200 0.0440 0.0875 0.168 0.330 0.658 1.301 9 0.0011 0.0130 0.0212 0.0435 0.0851 0.169 0.338 0.663 1.296 10 0.0007 0.0134 0.0194 0.0426 0.0879 0.171 0.326 0.664 1.308 Avg. 0.0010 0.0138 0.0197 0.0433 0.0857 0.168 0.329 0.655 1.305 S.D. 0.0012 0.0017 0.0015 0.0014 0.0015 0.0020 0.0037 0.0071 0.0110 C.V.% N/A 12.2 7.4 3.3 1.7 1.2 1.1 1.1 0.8 *D etect i o n l i m i t： 50 n g/uL *Qua n tit a t i o n l i m i t ： 100 n g/uL JASCO INC. 28600 Mary's Cour t ， E a s to n ， M D 21601US A