Life sciences
Explore the extensive applications of Hidex products in the field of life sciences, from kinetic assays and biodistribution studies, to nucleic acid and protein quantification, contributing to advancements in drug development, preclinical research, and analytical accuracy.
Drug Metabolism and Pharmacokinetics in Drug Discovery
Pharmacokinetics and Drug Metabolism (PKDM) play a vital role in optimizing compounds, both in vitro and in vivo. In recent years, it has become increasingly crucial to investigate Absorption, Distribution, Metabolism, and Elimination (ADME) characteristics during the early stages of drug discovery candidate selection to prevent potential issues in later development.
These assays impose significant demands on the detection systems, requiring versatility, sensitivity, performance, a range of detection technologies, various accessories, and precise environmental control, among other factors.
Particularly, ADME/Toxicity, Thymidine uptake, Receptor-Ligand and GTP Binding, Kinase, enzymatic molecular interaction, and Chromatographic assays that separate reaction mixtures into their components (fraction analysis combined with HPLC) greatly benefit from the use of radioactive labels.
For processes like whole cell proliferation (Thymidine uptake) and Cell-membrane (Receptor-Ligand Binding), filtration methods are typically employed. Samples are harvested into filtermats or filter bottom plates, and filtermats are conveniently measured using radioactive detection equipped Sense readers, utilizing Hidex MaxiLight scintillation liquid and Filtermat adapters.
There are also microplates available on the market with solid scintillators applied onto the well surface. In this case the samples are dried into the wells, and no liquid cocktail is needed. These are typically used with small-molecular samples e.g. from HPLC runs (fraction analysis). However, these are less suitable for macromolecular samples (proteins, plasma, cell debris) as these don’t dry properly and tend to form complex masses with high self-absorption.
When using cocktail mix liquid assays, it is important to use microplates which are chemically resistant to the cocktails. Otherwise, the microplates may degrade or dissolve, leading to signal loss and, in the worst case, contamination of the instrument. Standard polystyrene plates, for example, may not be suitable for such applications.
Hidex offers the Sense Beta and Sense Beta Plus, which are designed to meet the demands of these various applications. The Sense Beta Plus is the first microplate reader to combine radiometric and non-radiometric detection technologies.
Biodistribution
When developing a drug, certain milestones are essential before its release to the market, particularly in assessing the drug's ADME properties and safety.
Drug Metabolism and Pharmacokinetics (DMPK) is a foundational discipline in drug development. It involves the examination of a drug compound's biotransformation and other pharmacokinetic properties to evaluate its safety.
Preclinical research involving small animals has been the gold standard for decades and remains a crucial criterion for determining the safety and efficacy of products before they undergo clinical testing. The primary goal of preclinical research is to establish the safety profile of a compound.
Pharmacokinetics focuses on studying a compound's concentration in the body over time, and it is linked to the processes of absorption, distribution, metabolism, and excretion (ADME) of a compound.
Animal PK studies serve as invaluable tools for screening compounds during the candidate optimization phase of drug discovery. These studies provide valuable insights into compounds, including exposure, area under the curve (AUC), clearance, and half-life, among other parameters. Proper PK studies can expedite the discovery process and ensure that only compounds with favorable PK profiles progress to clinical trials.
Biodistribution is a fundamental aspect of identifying target organs and predicting safety and efficacy, and it can be integrated into safety and pharmacology studies.
In biodistribution studies, tissue weight is critical for calculating the percentage of administered dosage uptake and biodistribution. This typically involves manual work with a traditional analytical balance, which can be a time-consuming process.
Hidex offers the Automatic Gamma Counter with an onboard balance ideal for biodistribution studies. The Hidex AMG solution frees up time, automatically saves results, and increases the traceability of sample data.
Nucleic acid quantification
DNA and RNA concentration assays using photometric methods require blank sample subtraction and sample purity determination by measuring optical density (OD) at 260 nm, 230 nm, 280 nm, and 320 nm wavelengths. Nucleic acids generally absorb ultraviolet light at 260 nm, while the other wavelengths are utilized to determine the blank and purity of the sample. The Beer–Lambert law is used to calculate the concentrations without the need for standard curves.
The ratio of the absorbance at 260 nm and 280 nm is used to assess the purity of nucleic acids. In addition, an examination of other wavelengths in the sample spectra may be useful; for example, absorption at 230 nm can be measured to determine contamination by phenolate ions, thiocyanates, and other organic compounds. Absorption at 330 nm and higher indicates the presence of particles in the solution.
Also, the physical sample path length needs to be known for the concentration calculation, either by specifying the dimensions of the sample vial or by measuring the OD in the red-light region above 900 nm, where there is minimal impact from nucleic acid, proteins, organic compounds, or other sample impurities.
An alternative method to assess DNA and RNA concentration is to tag the sample with a fluorescent tag, e.g., Ethidium bromide. The benefit of fluorescence quantitation is improved sensitivity over spectrophotometric methods. However, in this case, the sample volumes need to be large enough to fit a microplate format, and protein contamination must still be determined using photometric methods.
Protein quantification
Protein quantification assays are quite common and are utilized to determine various parameters such as blood serum total protein quantification, the albumin/globulin ratio, binding constants in general, and enzyme kinetics.
There are several photometric methods to measure protein concentration, many of which rely on the presence of tryptophan and tyrosine in the protein sequence. Absorbance at 280 nm is simple and straightforward, although it may not be very accurate. Bradford and BCA assays are considered more accurate but require a known set of standard curve samples for quantification.
In the standard Hidex Sense and Sense Beta Plus models, the complete absorbance spectrum from 220 nm to 1000 nm is always recorded in less than a second. This allows for DNA and RNA concentration calculations to be done with a single measurement. It is also very easy to optimize the exact wavelengths and bandwidths for any assay, even offline after the actual measurements have been taken.
Our life science solutions redefine precision and accuracy. From the Hidex Automatic Gamma Counter facilitating biodistribution studies to the comprehensive capabilities of the Hidex Sense and Hidex Sense Beta Plus models for nucleic acid and protein quantification, our devices are ideal instruments developed for drug development studies, preclinical research, and analytical quantification in the pursuit of scientific excellence.
