Application Notes

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  • AN001: Concentration effects on measurements of hydrodynamic radius for standard proteins (pdf, 0.68MB)

    This application note shows use of the ActiPix™ for determination of hydrodynamic radius of a protein. The method allows proteins to be used in their native form, without any labelling or denaturation. A plug of protein solution is injected into a fused silica capillary, driven through the capillary by application of pressure, and detected using UV area imaging as it passes windows at entrance to and exit from a loop in the capillary. The radius of the protein is determined by analysis of band broadening due to Taylor dispersion. The method is applicable over a wide concentration range and uses only nanolitres of sample.

  • AN002: Multi-compound electrophoretic assays for biocatalytic activity with the ActiPix™ D100 (pdf, 0.62MB)

    This application note describes how the ActiPix™ UV area imaging detector can be used to test biocatalyst substrate specificity towards a mixture of UV active compounds using a continuous engagement electrophoretically mediated microanalysis (EMMA) assay method.

  • AN003: Substrate specificity screening with UV area imaging detector (pdf, 0.86MB)

    This application note describes how the ActiPix™ UV area imaging detector can be used to test penicillinase (β-lactamase) active towards a mixture of UV active substrates using an electrophoretically mediated microanalysis (EMMA) assay method.1 A variety of bacteria produce the β-lactamase enzymes which catalyse the ring-opening of β-lactam-based antibiotics. Identifying those compounds that are resistant to this process is important as it is responsible for bacterial resistance to penicillin and other β-lactam antibiotics.

  • AN004: lab-on-capillary systems (pdf, 0.87MB)

    This application note describes how the ActiPix™ UV area detector can be used in a number of lab-on-capillary setups that combine various reaction, separation and detection steps. By assembling integrated systems from readily available components nanoscale reactions can be carried out that minimise the consumption of expensive materials.

  • AN005: Rapid sizing of quantum dots and nanoparticles (pdf, 1.44MB)

    Paraytec have developed a new approach to determining the hydrodynamic radius of species in solution. This application note is a proof of principle study to demonstrate the use of the ActiPix™ HT Nano-Sizing System for determination of hydrodynamic radii of quantum dot and gold nanoparticle samples

  • AN006: Rapid Sizing of Proteins and Antibodies (pdf, 1.37MB)

    Paraytec have developed a new approach to determining the hydrodynamic radius of species in solution. This application note shows the applicability of the ActiPix™ TDA200 HT Nano-Sizing System for determination of hydrodynamic radii of protein and antibody samples.

  • AN010: Real-time measurement of Ca release from bovine tooth sample (pdf, 0.87MB)

    Paraytec have developed a new approach for real-time measurement of the amount of Ca2+ released from the surface of a bovine tooth sample. This technique is significantly faster and more sensitive than other current techniques. Great potential exists for quick measurement of anti-erosion consumer products.

  • AN011: Real-time measurement of nicotine release from dermal patch (pdf, 0.82MB)

    Paraytec have developed a new approach to measuring the amount of nicotine released from a sustained release dermal patch. Rate of release and insight into membrane transport information are explored.

  • AN012: Rapid determination of intrinsic dissolution rates of APIs using Surface Dissolution Imaging (pdf, 0.68MB)

    Surface dissolution imaging offers a rapid means of determining intrinsic dissolution rates (IDRs) of active pharmaceuticals ingredients (APIs). In this study the IDR of griseofulvin was determined in less than 20 minutes.

  • AN013: Analysis of peptides in nanoLC (pdf, 0.92MB)

    This application note illustrates use of the ActiPix™ D100 for detection of three peptides (angiotensin II, Met-enkephalin and Leu-enkephalin) following separation by nanoLC on a 75 mm i.d. column. Using a nanoLC cartridge with detection capillary having i.d. 75 mm, results are shown for loadings of 0.5 ng each peptide. The S/N for Leu-enkephalin is 63, suggesting that the limit of detection is below 0.05 ng (80 fmol). Detection is also demonstrated in the 20 mm i.d. fused silica transfer line, and with loading of 1 ng each peptide on column the S/N for Leu-enkephalin is 4.4. Peak broadening due to Taylor dispersion in the wider bore capillary causes the peaks to be slightly wider than those measured directly in the transfer line.

  • AN014: On column detection monolithic fused silica capillary (pdf, 0.68MB)

    This application note illustrates on-column detection using a monolithic capillary column 100 mm i.d./ 360 mm o.d. and isocratic elution. Flow resistance in monolithic columns is much lower than in conventional particle packed columns, and a syringe pump is used to drive the flow. For 1 pmol loading of caffeine and 0.5 s time constant, the signal-to-noise ratio is 23. This shows that the ActiPix™ D100 detector has femtomole sensitivity for on-column detection. A particular benefit of imaging through the column bed with the ActiPix™ D100 is that time-displaced averaging used in Paraytec’s proprietary software removes any random fluctuations in the signal due to non-uniformity of the stationary phase and scattering effects. As shown with a sharply fronting peak of caffeine under overload conditions, this occurs without any sacrifice of the 70 mm spatial resolution.

  • AN015: Precision and accuracy of protein sizing (pdf, 1.54MB)

    This application note outlines results with a test protein, myoglobin, using a technique developed by Paraytec for determining the hydrodynamic radius of species in solution. Repeatability and intermediate precision are reported, and the key factor influencing the accuracy of the method is documented. The technique measures the extent of Taylor dispersion of a plug of sample moving through a buffer solution. The extent of dispersion correlates directly with the hydrodynamic radius of the species.

  • AN016: Separation, size and charge determination of small molecules using the ActiPix™ D100 de (pdf, 1.14MB)

    This application note demonstrates integration of the Paraytec ActiPix™ D100 detector with an Agilent 7100 CE system to determine size, mobility and charge of components in a mixture of small molecules. The analytes lidocaine, phenylmethanol and benzoate are chosen as representative of small molecules of different charge type: cationic, neutral and anionic respectively in the background electrolyte used, phosphate buffer at pH 7.5. A capillary with three windows is used, with detection at the 1st window using the Agilent 7100 and at the 2nd and 3rd using the ActiPix™ D100. The separated species are characterised at the 1st window after short-end injection followed by pressure assisted capillary electrophoresis for 3.5 minutes. The hydrodynamic radii of the molecules are determined from the broadening of the bands between the 2nd and 3rd windows during a 15 minute stage of pressure driven flow. The method allows radii to be measured within a single run, with good repeatability (RSDs better than 2.5%, n=9). Charge follows by combining data on radius and mobility.

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