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Cell Line Optimization

Quantitation of Nucleic Acids BioSpec-nano

Quantitation of Double-Stranded benefits

DNA Using BioSpec-nano Cell Line Optimization
click here • Measure the concentration or check the purity of double-stranded DNA extracts.
• Measure sample quantities as small as 1 μL.
Operating Principle and Features Given the steps involved in one set, including measuring double- • Automatic wiping function enables a low-carryover system.
stranded DNA → wiping → adding TE buffer → wiping, repeating that
The BioSpec-nano has two available optical path lengths, 0.2 mm and set 60 times resulted in carryover (%) that remained 0.3 % or less,
0.7 mm, which enable quantitation of nucleic acids in very low sample which confirmed that sample carryover in the sample area when using
volumes of 1 or 2 μL. Samples can also be measured using an optional Culture
cell with a 5 mm optical path length (for 2 mL volumes of dilute automatic wiping is extremely low. With the automatic wiping function, never
samples). Table 1 Analytical Conditions forget to wipe off samples.
An automatic wiping function enables wiping the samples between Sample concentration 50 to 3700 ng/µL
measurements, eliminating the need to manually clean the sample stage Pathlength 0.2 mm Sample volume 1 µL
and reducing cross contamination between samples. Sample concentration 15 to 1000 ng/µL
Pathlength 0.7 mm
Sample volume 2 µL
Measurement Method 5 mm Pathlength Cell Sample concentration 2 to 150 ng/µL
(option) Sample volume 2 mL
The sample consisted of purified dsDNA dissolved in Tris-EDTA (TE) buffer
solution. The individual samples were prepared in the concentration ranges
listed in Table 1 for each pathlength. Next, 10 successive measurements Purification
were conducted using each of the pathlengths and concentrations using
the BioSpec-nano, and the OD (Optical Density, absorbance corresponding
to the 10 mm pathlength) at 260 nm was determined. The Y-axis values
(Measured OD260) in Fig. 1, 2, and 3 correspond to BioSpec-nano
measurement values. The standard value (Corrected OD260, X-axis in each
figure) for determining the accuracy was obtained using the Shimadzu
Ultraviolet-Visible spectrophotometer, an appropriately diluted sample
and a 1 mm pathlength cell. The linearities of Fig. 1, 2 and 3 indicate Fig. 1 Analysis Results with 0.2 mm Pathlength
the linearity of the standard values, and the deviation from each of the
straight lines correspond to OD error.
Results Characterization

Analysis Results with 0.2 mm Pathlength Specifications
The correlation coefficient of 0.999 for OD260 was obtained with
respect to the standard value (Fig. 1). When the OD value was greater Instrument BioSpec-nano
than 5 (250 ng/μL dsDNA), the measurement repeatability as CV (%)
was less than 1.4 %, and the OD error (%) was from -5.4 % to 2.8 %. Wavelength range 220 to 800 nm
The data are shown in Fig. 1. Fig. 2 Analysis Results with 0.7 mm Pathlength Spectrum bandwidth 3 nm
Analysis Results with 0.7 mm Pathlength Wavelength accuracy ±1 nm Quality Control
The correlation coefficient of 0.999 for OD260 was obtained with Pathlength 0.2 mm, 0.7 mm
respect to the standard value (Fig. 2). When the OD value was greater Photometric value unit OD (Optical Density), absorbance converted with 10 mm pathlength
than 1.4 (70 ng/μL dsDNA), the measurement repeatability as CV (%)
was less than 1.4 %, and the OD error (%) was from -8.6 % to 4.4 %. Sample volume 1 µL min. (pathlength: 0.2 mm)
The data are shown in Fig. 2. 2 µL min. (pathlength: 0.7 mm)
Light source Xenon flash lamp
Analysis Results with 5 mm Pathlength Cell
The correlation coefficient of 0.999 for OD260 was obtained with Monochromator Holographic grating
respect to the standard value (Fig. 3). When the OD value was greater Fig. 3 Analysis Results with Optional 5 mm Pathlength Cell Detector Photo diode array
than 0.2 (70 ng/μL dsDNA), the measurement repeatability as CV (%)
was less than 0.6 %, and the OD error (%) was from -1.6 % to 3.6 %. Summary Auto wiping function Provided Pharmacokinetics
The data are shown in Fig. 3. Spectrum measuring time 3 sec
BioSpec-nano is capable of simple and excellent measurement linearity,
Performance of Automatic Wiping in Nucleic Acid Quantitation reproducibility, and accuracy with a sample volume of 1 to 2 μL for Quantitation range Pathlength 0.2 mm, 1 to 75 OD, 50 to 3,700 ng/µL
We alternated measurement of purified dsDNA (11.7 OD, 578 ng/ optical pathlengths of 0.2 mm and 0.7 mm, respectively. Pathlength 0.7 mm, 0.3 to 21 OD, 15 to 1,000 ng/µL
Optional 5 mm pathlength cell, 0.04 to 3 OD
μL) and TE buffer solution using a 0.7 mm pathlength, 3 μL sample
volume, and 1 wipe operation between measurements. Carryover Application Examples Dimensions W 210 mm × D 214 mm × H 417 mm
(%) of dsDNA to the TE buffer solution was used as an index of the
automatic wiping performance. • Measuring single-strand DNA concentration Weight 7 kg Others
• Measuring RNA concentrations Analysis mode Simple nucleic acid quantitation, labeled nucleic acid quantitation, protein quantitation, labeled
Carryover (%) • Measuring protein concentration (refer to p. 34) protein quantitation, photometric measurement
[(Nucleic acid concentration in TE measurement)] Note: The droplet formation status will affect analysis results. Measure quantities that are large enough to enable proper droplet formation.
= 100× … (1)
[(Nucleic acid concentration in dsDNA measurement)]

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