Nucleoside Nucleotide Quantification
Nucleoside Nucleotide Quantification - Related Videos
06:38
Plant Sample Preparation for Nucleoside/Nucleotide Content Measurement with An HPLC-MS/MS
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Cited by 3 •
2021
A precise and reproducible method for in vivo nucleosides/nucleotides quantification in plants is described here. This method employs an HPLC-MS/MS.
04:32
Chip-in-a-Tube-Based Digital PCR for Quantification of Single Nucleotide Variants
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2025
This video demonstrates the utility of digital PCR in a chip-in-a-tube format for the detection of single nucleotide variations in DNA samples. A single PCR reaction is partitioned into chambers that act as independent PCR reactions, and the detection of fluorescence signals from the amplified targets in the chambers is used to compute the frequency of the variant allele in the sample.
11:29
HPLC-based Assay to Monitor Extracellular Nucleotide/Nucleoside Metabolism in Human Chronic Lymphocytic Leukemia Cells
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Cited by 5 •
2016
The protocol described here represents an easy and reproducible method that employs reverse phase high-performance liquid chromatography (RP-HPLC) to measure purine metabolism on chronic lymphocytic leukemia (CLL) cells cultured under different conditions.
15:22
Nucleoside Triphosphates - From Synthesis to Biochemical Characterization
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Cited by 4 •
2014
The protocol described herein aims to explain and abridge the numerous obstacles in the way of the intricate route leading to modified nucleoside triphosphates. Consequently, this protocol facilitates both the synthesis of these activated building-blocks and their availability for practical applications.
02:48
Labeling of Replicating HSV-1 Genomes with a Nucleoside Analog
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2026
Source: Dembowski, J. A., Deluca, N. A. Purification of Viral DNA for the Identification of Associated Viral and Cellular Proteins. J. Vis. Exp. (2017)This video demonstrates the infection of stationary-phase mammalian cells with HSV-1 (Herpes Simplex Virus-1), followed by replication of the viral genome using host and viral replication machinery. A synthetic nucleoside analog is incorporated into newly synthesized viral DNA to label replicating genomes, which are subsequently isolated for...
FAQs
Chromatographic separation matters because biological extracts contain multiple polar metabolites whose signals may otherwise be difficult to distinguish. High-performance liquid chromatography separates these compounds before ultraviolet detection or mass spectrometry, creating a clearer basis for identifying and measuring individual nucleosides and nucleotides. This separation supports more reliable interpretation of changes in energy status and nucleic acid metabolism.
Calibration standards provide the reference needed to relate an analytical signal to the amount of a target compound. Internal standards help account for variation introduced during extraction and measurement. Used together, they strengthen quantitative comparisons among samples and reduce the risk that differences in handling or instrument response will be mistaken for genuine biochemical changes.
Some nucleosides and nucleotides are unstable, so delays or unsuitable extraction conditions can alter their measured abundance before analysis. Careful sample handling preserves the original metabolite composition, while controlled extraction prepares the polar compounds for chromatographic separation. These safeguards are especially important when comparing cellular stress, drug responses, or genetic changes across experimental samples.
A typical workflow begins by collecting and extracting the biological or biochemical sample under controlled conditions. The extract is then separated by high-performance liquid chromatography, followed by ultraviolet detection or mass spectrometry for identification or measurement. Calibration standards establish the quantitative reference, while internal standards and consistent handling improve confidence in the resulting values.
Researchers can apply these measurements to examine ATP and other energy carriers, assess activity in nucleic acid metabolism, and investigate biochemical pathway changes. The approach also supports studies of enzyme function, cellular stress, and responses to drugs or genetic changes. Comparing metabolite measurements across conditions can therefore connect molecular changes with broader cellular behavior.
Measured nucleoside and nucleotide levels can provide evidence about cellular energy status, pathway activity, and the consequences of altered enzyme function. In experimental systems, shifts in these compounds may help characterize stress responses or changes caused by treatment and genetic manipulation. The results are most informative when sample preparation, standards, and analytical conditions remain consistent across comparisons.