Report Example on Ion Exchange Chromatography

Introduction

Columns packed with ion exchanger aid in the ion exchange separations—commercially available ion exchangers of styrene and benzene. For instance, the DEAL cellulose is a good example. The charge of the particles to be separated determine the choice of the exchanger. Hence anion exchanger separates anions while a cation exchanger separates cations (Zhang et al.,2019). Procedurally the ion exchanger is the first to be placed in the column, followed by the sample to be separated, and then lastly, a buffer solution. The most commonly used buffers include tris-buffer, pyridine buffer, acetate buffer, and phosphate buffer. Affinity for the ion exchanger separates the particles allowing those with high affinity to move along with the buffer. The particles are later analyzed spectroscopically after the separation of tightly bound buffers.

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Materials and Methods

The DNA segment amplification is by the polymerase chain reaction that is entirely biochemical and in vitro. The procedure involves three vital steps, and these include denaturing, annealing, and elongation. In the DNA denaturing, the weak hydrogen bonds are broken through heating at 940 C and hence the separation of the double strands into single strands (Rischawy et al.,2019). In the template DNA, the primers bind the complementary sequence at a temperature of 50-70o C in the annealing stage. In the DNA elongation stage, the DNA polymerase adds nucleotides to the primer sequentially with a DNA template. The optimal working temperature is 72 0 C and, therefore, the temperature for DNA extension. The result for one cycle is two different separate double-stranded DNA that are templates in the next subsequent cycles. The production of these copies of DNA is exponential throughout the repeated cycles.

Reverse Phase Chromatography

the instrument condition should be verified before the onset of the experiment. To prevent silica from collapsing, water in the mobile phase is vital. The procedure requires two different solvents. In any reverse chromatography, the setting of the rate of flow, pressure, type of column used, detection parameters such as wavelengths when using ultraviolet light determine its effectiveness. The sample to be measured is first dissolved in a suitable solvent, for instance, methanol. Centrifugation removes massive particles and later to collect the filtrate using a filter paper. The sample is ready for loading into the vial.

Fluorescence Spectroscopy

The substances' basic principle involves absorbing an appropriate wavelength's exciting radiation and emitting the light appropriately. A signal generated after detecting the radiation is directed to a surface of interest and is compared to an acceptable standard, and has high cleanliness levels (Nambiar et al.,2018). Manual or automatic focusing of the different locations on a single spectrum helps achieve high spatial resolution. Besides assessing for contamination in the healthcare institutions, the procedure helps in assessing dermal exposure to contaminants.

Methodology

More energetically stable atomic nuclei are formed from the disintegration of unstable atomic nuclei. The characterization depends on the initial large nuclei's sample half-life since it is a first-order process. The process is highly exothermic, liberating more energy than an exothermic chemical reaction. Changes in temperature and pressure have no significant effects on this process since it is a nuclear phenomenon (Reyes et al.,2017). It is important to balance nuclear reactions and equations by labeling the unstable species with mass numbers and atomic numbers.

Densitometry

The ability to measure optical density in light-sensitive materials such as photographic film when exposed to light is densitometry. Densitometry measures bone mineral density. The procedure involves a low amount of radiation exposure in addition to the soft skills required to perform it (Swinney et al.,2018). Two different beams, one with high energy and another with low energy, are produced, measuring the amount of radiation passing the bone. Bone density is measured following the differences in the two beams caused by bone thickness variations (Jeong et al.,2018). The main body parts involved in densitometry are the spine, hip, and forearm. During the test, the patient lies on the table for approximately 10-20 minutes. The process is painless, and no injections are used for testing.

Conclusion

Various methods are used in the labeling of the nucleic acids depending on the desired application. Nucleic acid labeling with enzymes, radioactive phosphates, and fluorophores is done through enzymatic and chemical methods. Highly specific probes with the label distributed through the nucleic acid are vital for hybridization reaction such as southern and northern blots (Shi et al.,2018). End-labeled probes are essential in applications involving protein interactions such as gel-shift assays. In detecting specific DNA/RNA sequences in a complex mixture of nucleic acids in southern blot, northern blot, and situ, hybridization between immobilized and labeled probes is essential. The nature of the probe determines the specific detection methods.

References

Jeong, L. N., & Rutan, S. C. (2018). Simulation of elution profiles in liquid chromatography–III. Stationary phase gradients. Journal of Chromatography A, 1564, 128-136.

Nambiar, A. P., Sanyal, M., & Shrivastav, P. S. (2018). Simultaneous densitometric determination of eight food colors and four sweeteners in candies, jellies, beverages, and pharmaceuticals by normal-phase high-performance thin-layer chromatography using a single elution protocol. Journal of Chromatography A, 1572, 152-161.

Reyes, B., Coca, M. I., Codinach, M., López-Lucas, M. D., del Mazo-Barbara, A., Caminal, M., ... & Moraleda, J. M. (2017). Assessment of biodistribution using mesenchymal stromal cells: Algorithm for study design and challenges in detection methodologies. Cytotherapy, 19(9), 1060-1069.

Rischawy, F., Saleh, D., Hahn, T., Oelmeier, S., Spitz, J., & Kluters, S. (2019). Good modeling practice for industrial chromatography: mechanistic modeling of ion-exchange chromatography of a bispecific antibody. Computers & Chemical Engineering, 130, 106532.

Shi, M. T., Yang, X., A., & Zhang, W. B. (2019). Magnetic graphitic carbon nitride nano-composite for ultrasound-assisted dispersive micro-solid-phase extraction of Hg (II) before quantitation by atomic fluorescence spectroscopy. Analytica Chimica Acta, 1074, 33-42.

Swinney, M. W., Peplow, D. E., Patton, B. W., Nicholson, A. D., Archer, D. E., & Willis, M. J. (2018). A methodology for determining the concentration of naturally occurring radioactive materials in an urban environment. Nuclear Technology, 203(3), 325-335.

Zhang, Z., Zhou, S., Han, L., Zhang, Q., & Pritts, W. A. (2019, August). Impact of linker-drug on ion-exchange chromatography separation of antibody-drug conjugates. In MAbs (Vol. 11, No. 6, pp. 1113-1121). Taylor & Francis.