Per- and polyfluoroalkyl substances (PFAS) are synthetic chemicals that have been widely used since the 1940s in industries and consumer goods. They are found in products like water-repellent clothing, stain-resistant fabrics, cosmetics, firefighting foams, and grease-, water-, and oil-resistant items. For instance, PFAS are commonly present in non-stick cookware used in everyday life. Their persistence and long biological half-lives have resulted in measurable levels in nearly all people in developed countries, with associated PFAS human health effects reported worldwide.Evaluating PFAS health risks requires understanding their adverse health effects at real-world exposures, particularly during sensitive life stages, along with their modes of action, toxicokinetics, and dose-response relationships. This knowledge supports risk estimates that guide public health limits, exposure reduction, and environmental cleanup efforts.<h3>Why are PFAS a Health Concern?</h3>PFAS have a chemical structure with a perfluorinated carbon tail that repels both water and oil and an anionic head group that attracts water, making them highly resistant and persistent. As a result, they do not break down and instead accumulate in the bodies of people and animals worldwide through food and repeated exposures. They even bio-magnify through the food web. Growing evidence has raised concerns about the harmful effects of PFAS on human health.In the human body, PFAS bind strongly to proteins due to their polar, hydrophobic nature and have been detected in serum, cord blood, and breast milk. Unlike many other bio-accumulative contaminants, they do not tend to accumulate in fat tissue.<h3>PFAS Testing Techniques</h3>PFAS have a chemical structure with a perfluorinated carbon tail that repels both water and oil and an anionic head group that attracts water, making them highly resistant and persistent. As a result, they do not break down and instead accumulate in the bodies of people and animals worldwide through food and repeated exposures. They even bio-magnify through the food web. Growing evidence has raised concerns about the harmful effects of PFAS on human health.In the human body, PFAS bind strongly to proteins due to their polar, hydrophobic nature and have been detected in serum, cord blood, and breast milk. Unlike many other bio-accumulative contaminants, they do not tend to accumulate in fat tissue.While animal studies show that most PFAS are primarily excreted through urine, renal clearance in humans is much slower, raising significant concerns for PFAS human health. The slow elimination contributes to the long-term effects of PFAS on humans.PFAS have extended half-lives, with perfluorooctanoic acid (PFOA) persisting for approximately 3.5 years and perfluorooctane sulfonic acid (PFOS) for roughly 4.8 years. Some studies also report a range of 2.3 to 3.3 years. These findings highlight serious PFAS health risks, demanding the need for continued monitoring and risk assessment.<h3>Known and Suspected Health Effects of PFAs in Humans</h3><h4>Immunological effects</h4>Epidemiological studies have shown that PFAS exposure is linked to weakened immune responses, including reduced antibody levels after vaccinations such as diphtheria, tetanus, rubella, and mumps, with these effects persisting into adolescence and adulthood.Prenatal and early-life exposure to PFAS has been linked to higher risks of infections such as airway, throat, and gastrointestinal illnesses, as well as conditions like atopic dermatitis. Overall, the evidence on PFAS and human health suggests that exposure to PFAS during infancy and childhood can suppress the immune system, though some effects may differ by sex.<h4>Liver disease and cancer</h4>The liver is a major site for long-chain PFAS accumulation, with experimental studies showing toxic effects such as fat infiltration in liver cells, apoptosis, disrupted fatty acid metabolism, and even tumor development. Animal toxicology and histology studies indicate that PFAS can alter liver metabolism, causing increased bile acid reuptake and lipid buildup. These findings demonstrate the effects of PFAS on liver function and long-term disease risk.<h4>Kidney diseases</h4>PFAS exposure has been consistently linked to higher uric acid levels, a biomarker of increased risk for kidney disease, in both adults and children. The prolonged half-lives of long-chain PFAS are largely due to strong kidney reabsorption. Human studies link legacy PFAS such as PFOA and PFOS exposure to reduced kidney function and chronic kidney disease. PFOA and PFOS health effects can also be seen in the body where these substances tend to accumulate in kidney tissues causing nephrotoxicity.<h4>Reproductive outcomes</h4>Exposure to PFOA has been linked to reduced sperm motility, lower sperm concentration and count, and hormonal changes in young men. PFAS can also cross the placenta and accumulate in breast milk, allowing transfer from mother to child. As a result, young children often have higher serum PFAS levels than their mothers.<h3>How do PFAS enter the Environment and Food Chain?</h3>Human exposure to PFAS occurs through multiple pathways. These may vary from exposure to indoor air and dust to ingestion of contaminated water and food, particularly in areas near industrial sites. Thus, PFAS human health concerns may arise from:<ul><li>Water: PFAS contaminate public and private drinking water sources, rivers, and groundwater near industrial areas, landfills, and waste sites. Firefighting foams (AFFFs) used at airports, military bases, and training facilities are a major contributor, raising concerns about PFAS chemicals health effects on the communities dependent on these resources.</li><li>Soil and Air: They spread through soil and air around hazardous waste sites, manufacturing facilities such as chrome plating, electronics, textiles, paper, and through biosolids used as fertilizers.</li><li>Food: PFAS enter the food chain through contaminated fish, seafood, dairy, meat, crops, and animal products. Food packaging such as grease-resistant paper, fast-food wrappers, and microwave popcorn bags also contribute PFAS to the environment.</li><li>Household and Consumer Products: Everyday items like stain- and water-resistant fabrics, non-stick cookware, cleaning agents, paints, sealants, and certain personal care products (shampoo, dental floss, cosmetics) release PFAS into homes.</li><li>Exposure Pathways: This is an area of growing research, it is currently understood that people are being exposed to PFAS through various means, including drinking contaminated water, eating contaminated food, inhaling polluted air or dust, swallowing soil, using PFAS-containing products, or working in high-risk occupations such as firefighting and chemical manufacturing.</li></ul><h3>Importance of Accurate PFAS Testing</h3>Accurate<a target="_blank" rel="noopener noreferrer" href="https://www.phenomenex.com/industries/pfas"> PFAS testing solutions</a> are essential to measure the presence of environmental chemicals and metabolites in human tissues and fluids. Testing can provide a clearer picture of total internal exposure from all sources and pathways. The wide diversity of PFAS, with varying chemical and physical properties, requires the development of tailored analytical procedures that depend on the sample type and medium.The choice of biological sample depends on the specific PFAS of concern, with serum most often used to measure those with long half-lives, while urine or whole blood may be better for short-lived compounds. PFAS can also be detected in plasma, breast milk, and hair, each providing unique insights into exposure at different life stages. For example, hair can reflect historical exposures, and breast milk testing helps estimate infants intake, which is a significant concern for many communities that have been exposed to these substances.Because PFAS detection limits are 1,000 times lower (ppt) than routine analyses (ppb), strict precautions such as verifying PFAS-free supplies are essential to prevent sample contamination, which can still occur from vendor or manufacturing changes. Extremely low detection limits and regulatory criteria is in the parts per trillion (ppt) range. Hence, field QC sample collection and analysis are crucial in investigating the effects of PFAS on humans.

Health Risks Associated with PFAS Exposure

<div><div>Scientists from UCT Prague contributed to a large-scale study showing that a small change in the so-called anticodon stem of transfer RNAs can expand their decoding capabilities – not only in eukaryotes but also in bacteria.</div><div>Scientists from UCT Prague contributed to a large-scale study showing that even a small change in transfer RNA (tRNA) can fundamentally affect how a cell reads its genetic information. Computer simulations explained why certain tRNAs are able to ignore a signal that under normal circumstances marks the end of protein synthesis. Such tRNAs were previously known in higher organisms. The study, however, mapped how common these unusual tRNAs are in bacteria.The genetic code is a&nbsp;set of rules by which cells translate genetic information during protein synthesis. Transfer RNAs (tRNAs) play a&nbsp;key role in this process, recognising short stretches of mRNA known as codons through their anticodon. In addition to amino acid codons, there is one start codon and several termination (stop) codons. Stop codons are not recognised by transfer RNAs but by specialised enzymes that terminate protein synthesis. Standard tRNAs have five nucleobase pairs in their so-called anticodon stem. Recently, however, it has emerged that some organisms carry functional tRNAs with only four pairs. These "shortened" variants can recognise even codons that, according to the classical rules, should not correspond to them.An international team led by the Biology Centre of the Czech Academy of Sciences in České Budějovice analysed over 42,000&nbsp;bacterial genomes and found that tRNAs with a&nbsp;four-pair stem occur in bacteria surprisingly often. In some cases, these variants likely enable the reading of the stop codon UGA as the amino acid tryptophan, a&nbsp;phenomenon previously associated mainly with unicellular eukaryotes.The <a target="_blank" rel="noopener noreferrer" href="https://kolar.vscht.cz/">Kolář group</a> at the Department of Physical Chemistry, UCT Prague, focused on the question of why a&nbsp;four-pair stem alters the decoding properties of tRNA. To address this, they employed atomistic molecular dynamics simulations, which make it possible to track the movement of every atom in a&nbsp;tRNA molecule over time.Jakub Žváček, a&nbsp;student on the Master's&nbsp;programme in Data Engineering in Chemistry, prepared a&nbsp;total of nine variants derived from the tryptophan tRNA of the bacterium Escherichia coli: the wild type with a&nbsp;five-pair stem, six mutants with a&nbsp;disrupted top base pair, and two previously described variants that promote reading of the stop codon UGA (the G24A&nbsp;and A9C&nbsp;mutations). The simulation results showed that disrupting the top base pair does not cause the overall three-dimensional structure of the tRNA to collapse. It remains stabilised by stacking interactions of neighbouring bases and non-canonical hydrogen bonds. The key finding, however, concerns a&nbsp;region at the interface of the D-arm and the stem – kind of hinge within the tRNA. In some mutants, the simulations revealed destabilisation of this region and increased flexibility of the anticodon stem, reminiscent of the behaviour of known mutations that promote stop codon readthrough. The simulations thus provided a&nbsp;structural explanation for the experimental observations and helped formulate predictions about which nucleotide combinations in the disrupted pair are functionally relevant.The study proposed an extension of the so-called superwobble hypothesis to include non-canonical C:A&nbsp;pairing at the third codon position. If this hypothesis is confirmed for other types of tRNA as well, it will have profound implications for our understanding of how the genetic code — shared by all known organisms — has been shaped by evolution.The results were published in the <a target="_blank" rel="noopener noreferrer" href="https://academic.oup.com/nar/article/54/7/gkag327/8659125">Nucleic Acids Research</a> (2026, 54, gkag327).</div><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/VSCHT_Praha_Pocitacove_simulace_osvetlily_jak_neobvykle_t_RNA_meni_pravidla_cteni_genetickeho_kodu_f6de6700a0.jpg" alt="VŠCHT Prague: Computer simulations shed light on how unusual tRNAs change the rules of reading the genetic code: Molecular architecture of the transfer RNA with the 5th anticodon-stem base pair labelled. This base pair affects the decoding ability of the tRNA anticodon." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_VSCHT_Praha_Pocitacove_simulace_osvetlily_jak_neobvykle_t_RNA_meni_pravidla_cteni_genetickeho_kodu_f6de6700a0.jpg 245w," sizes="100vw" width="759" height="268"><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/VSCHT_Praha_Pocitacove_simulace_osvetlily_jak_neobvykle_t_RNA_meni_pravidla_cteni_genetickeho_kodu_2_c4296b9015.jpg" alt="VŠCHT Prague: Computer simulations shed light on how unusual tRNAs change the rules of reading the genetic code: Analysis of anticodon stem flexibility showed that the mutation of the 5th nucleobase pair (C27A), which enables stop-codon readthrough, loosens the anticodon stem in a manner similar to the previously known Hirsh mutation G24A. Conversely, the C27U mutation does not loosen the stem, which correlates with the absence of this mutation in the genomes of bacteria containing tRNAs with 4bp stems." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_VSCHT_Praha_Pocitacove_simulace_osvetlily_jak_neobvykle_t_RNA_meni_pravidla_cteni_genetickeho_kodu_2_c4296b9015.jpg 245w," sizes="100vw" width="759" height="196"><blockquote><h4>Original article&nbsp;</h4><h4><a target="_blank" rel="noopener noreferrer" href="https://academic.oup.com/nar/article/54/7/gkag327/8659125?login=false">Frequent occurrence and predicted functions of tRNAs with 4-base-pair anticodon stems in bacteria: extended superwobble hypothesis</a></h4>Fadel Fakih, Satish Nandipati, Ambar Kachale, Jiří Heller, Jakub Žváček, Filip Brázdovič, Nawal Al-Chamy, Pragya Tripathi, Zdeněk Paris, Leoš Shivaya Valášek, Michal H Kolář, Vyacheslav Yurchenko, Marek Eliáš, Eugene V Koonin, Julius Lukeš, Anzhelika ButenkoNucleic Acids Research, Volume 54, Issue 7, 24 April 2026, gkag327<a target="_blank" rel="noopener noreferrer" href="https://doi.org/10.1093/nar/gkag327">https://doi.org/10.1093/nar/gkag327</a><h6>licenced by <a target="_blank" rel="noopener noreferrer" href="https://creativecommons.org/licenses/by/4.0/">CC-BY 4.0</a></h6></blockquote><h4>Abstract</h4>Recently, a tRNATrpCCA with a 4-base-pair (bp) anticodon stem (AS) was shown to efficiently recognize a near-cognate UGA codon in unicellular eukaryotes, such as some trypanosomatids and ciliates, thereby representing a novel codon reassignment mechanism. To determine whether this mechanism also evolved in bacteria, we analysed a dataset of 42 109 genomes, including previously reported cases of stop-to-tryptophan UGA reassignment and a newly identified instance in the phylum Patescibacteriota. We show that the 4-bp AS tRNATrp species are present across diverse bacteria and in some cases likely function in decoding in-frame UGA codons. Most notable is the endosymbiotic bacterium Candidatus Zinderia insecticola, which contains only the near-cognate 4-bp AS tRNATrpCCA, while lacking both canonical 5-bp AS tRNATrpCCA and a tRNATrpUCA. The secondary structure of this 4-bp AS tRNATrp resembles that of its eukaryotic counterpart, suggesting convergent evolution. We experimentally confirmed the UGA readthrough capacity of 4-bp AS tRNATrpCCA in Escherichia coli, and applied molecular dynamics simulations to suggest the underlying mechanism. Furthermore, we tested several predictions based on accepting the previously excluded possibility of C:A base pairing at the 3rd codon position. These findings provide new insights into the structural diversity of transfer RNAs (tRNAs) and expand our understanding of genetic code evolution.</div>

Computer simulations shed light on how unusual tRNAs change the rules of reading the genetic code

Clarity 10.2 expands instrument control capabilities with new control modules and updates the MS extension to help you work more efficiently.<h4><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/999">Clarity</a> 10.2 highlights</h4><ul><li>New and updated control modules</li><li>Improved MS data import and export</li><li>Obsolete hardware support ended</li></ul><h4>Instrument control modules</h4>Clarity 10.2 introduces new instrument control modules from Annuo (AS-3117, FAS-3823) and updated instrument controls form Sykam and Ecom.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Data_Apex_Instrument_control_modules_5de26782b0.jpg" alt="DataApex: Instrument control modules" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Data_Apex_Instrument_control_modules_5de26782b0.jpg 228w," sizes="100vw" width="380" height="260"><h4>MS data import/export improvements</h4>Clarity improves MS data exchange, especially for *.cdf files. MS instruments now support export and import of multiple spectral data and non-spectral signal data in the same CDF file. Import also better handles uniform sampling information, m/z Step values, scale factors, and file names with non-ASCII characters. Exporting individual chromatogram signals from MS instruments is now available through a separate Export Chromatogram dialog.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Data_Apex_MS_data_import_export_improvements_a7967a4020.jpg" alt="DataApex: MS data import-export improvements" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Data_Apex_MS_data_import_export_improvements_a7967a4020.jpg 228w," sizes="100vw" width="378" height="259"><h4>Obsolete hardware support terminated</h4>Due to changes in Windows driver security requirements, this update discontinues support for obsolete hardware, including INT7, INT9, U-PAD, U-PAD2, and CB20.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Data_Apex_Obsolete_hardware_support_terminated_24d9329419.jpg" alt="DataApex: Obsolete hardware support terminated" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Data_Apex_Obsolete_hardware_support_terminated_24d9329419.jpg 228w," sizes="100vw" width="378" height="259"><blockquote>Explore the complete list of all changes, fixes, and additions in version 10.2.👉<a target="_blank" rel="noopener noreferrer" href="https://www.dataapex.com/version_history?product=1">See the complete changelog&nbsp;</a></blockquote>

Clarity Chromatography Software 10.2 released!

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Science and research

Software

Since 1991 DataApex develops and manufactures chromatography data stations. DataApex is constantly striving to provide users with efficient and up-to-date tools for chromatography data processing.

DataApex

The University of Chemistry and Technology, Prague is a natural centre of first-rate study and research in the area of chemistry in Czechia and is one of the country's largest educational and research institutions focused on technical chemistry, chemical and biochemical technologies, material and chemical engineering, food chemistry, and environmental studies.

UCT Prague

Phenomenex is a global technology leader committed to developing novel analytical chemistry solutions that solve the separation and purification challenges of researchers in industrial, government and academic laboratories.

Phenomenex

Watrex xTrovert XT Automatic DNA/RNA extractor

Watrex xTrovert XT is an automatic DNA/RNA extractor. It streamlines the extraction process, making it more efficient and reproducible.

Automatic extractor utilises a simple principle of two pipetting heads serving a 96-well plate.Thanks to easy replacement head options of pipetting with 300 μl tips or 1000 μl tips are available. Automatic optical sensor calibration enables the use of pipetting tips and 96-well plates from virtually any supplier. The device is equipped with two magnetic arms for protocols including paramagnetic SPR beads. It also allows for thermal incubation in the range of 25 to 85 °C. In addition to internal LED lighting, the station is equipped with an integrated UV-C sterilisation unit. Intuitive control from TFT touch panel user interface facilitates easy set-up. xTrovert™ XT is a fully programmable open system where user can create their own workflow scripts downloadable as text files from USB flas disk.<h3>APPLICATIONS</h3><ul><li>Sample prep for PCR testing</li><li>NGS library prep</li><li>Sanger DNA sequencing</li></ul><h3>CURRENT PROTOCOLS</h3><ul><li>DNA/RNA extraction</li><li>DNA purification</li><li>PCR clean-up/desalting</li></ul><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Watrex_Watrex_x_Trovert_XT_Automatic_DNA_RNA_extractor_Compatible_with_any_magnetic_bead_based_kit_23886eea52.jpg" alt="Watrex: Watrex xTrovert XT Automatic DNA/RNA extractor - Compatible with any magnetic bead based kit"><h3>TECHNICAL SPECIFICATION</h3><ul><li>Capacity: 16 or 32 samples/run</li><li>Volume range: 10 – 600 μl</li><li>Temperature: 25 – 85 °C</li><li>Sterilization: UV-C</li><li>Display: 3,5“ TFT touch display</li><li>Connectivity: USB 1.0/2.0</li><li>Power source: 24 V adaptor</li><li>Dimensions in cm: 65(w) x 65(h) x 34(d)</li><li>Weight: 25 kg</li></ul><h3>Ordering information:</h3><h5>PN/Component:</h5><ul><li>XTR000-001 xTrovert™ XT Universal robotic pipettor for DNA/RNA extraction &amp; handling</li><li>XTR000-003 Replaceable 8-channel 1000uL pipetting head for xTrovert™ XT</li><li>XTR000-004 Replaceable 8-channel 300uL pipetting head for xTrovert™ XT</li><li>XTR000-100 Deepwell plates (empty) for xTrovert™ XT</li><li>XTR000-220 DNA/RNA extraction kit, mgn. beads, 16-rxns per plate</li></ul>The entire system was designed and developed by Watrex Praha and is manufactured in the Czech Republic and promises a shift in the performance and efficiency of laboratory processes worldwide!You can also visit this product on the <a target="_blank" rel="noopener noreferrer" href="https://watrex.com/obchod/laboratory-automation/xtrovert-xt/?utm_source=labrulez&amp;utm_medium=propagace&amp;utm_campaign=labrulez_produkty">Watrex website</a>.

Quantitative analysis of proteins (Bioanalytic) 

In our laboratory, proteomic analyses, including quantitative analysis of proteins, are performed by HPLC-MS or HPLC-MS/MS methods on a ZenoTOF 7600 high-resolution mass spectrometer (SCIEX).

Quantitative analysis of proteins is important in many industries and applications. Proteins include a diverse group of compounds that participate in virtually every biological process. Proteins are building blocks, hormones, transport molecules, cell receptors, cell growth and differentiation factors. Because of this, the analysis of protein interactions is crucial in drug design and production.Proteins are macromolecules - the basis of their structure are polypeptide chains containing from 100 to more than 10000 amino acids (10 - 100 kDa). Proteins are found in simple and complex forms. Complex proteins contain a non-protein component (sugar, lipids, metal ions or dyes) in their structure.In our laboratory, proteomic analyses, including quantitative analysis of proteins, are performed by HPLC-MS or HPLC-MS/MS methods on a <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?manufacturerItems=52&amp;search=ZenoTOF+7600">ZenoTOF 7600</a> high-resolution mass spectrometer (SCIEX). We process the obtained results using Sciex OS, Spectronaut, Skyline software.<h4>MS vs MS/MS</h4>Quantitative analysis of intact protein (whole protein in intact form) leads to the concentration (determined from a prepared curve from a standard (with or without matrix) and confirmation of the identification by superimposing (on a mirrored basis) the MS spectrum of the protein determined in the sample to the MS spectrum of the standard. For your needs, we can generate a list of protein-structured compounds (impurities) that were identified in the sample and determine the ratio of their signal area to the signal area of the analyzed protein.The result of proteomic quantitative peptide analysis allows precise and much more sensitive determination of protein concentration. The analysis is based on MS/MS data obtained for peptides obtained by digestion with proteolytic enzymes. The reported result consists of the concentration of the protein under study and confirmation of its presence based on the determined amino acid sequence. For such analyses, it is recommended to add standards internal standards.<blockquote>Our offer:<ul><li>For proteins and peptides with low masses - intact analysis with MS detection;</li><li>Proteins with masses &gt;20kDa require optimization of the digestion method and MS/MS detection. This results in specific signals from the analyzed protein in the mixture/matrix;</li><li>From complex matrices (including serum, cell culture, fungal suspension), sample analysis involves several steps:&nbsp;<ul><li>Sample purification;</li><li>Digestion to obtain unique peptides to distinguish the analyzed proteins from other proteins in the sample;</li><li>Sample preparation requires appropriate method optimization and validation (let's set validation criteria individually);</li></ul></li><li>Reported quantitative result includes confirmation of the presence of the protein in native form or a peptide derived from it (usually confirmation includes at least two peptide sequences);</li><li>Statistical analysis using tools: MarkerView (PCA), GraphPad Prism, Trends, Sciex OS.</li></ul>To optimize and validate the method, it is necessary to provide a standard, in the case of proteins with high masses - it is recommended to add an internal standard.See also:<ul><li><a target="_blank" rel="noopener noreferrer" href="https://labexperts.com.pl/service/identyfikacja-bialek/">Protein identification</a>,</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1296">Analysis and characterization of monoclonal antibodies</a>.</li></ul></blockquote><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Bioanalytic_Quantitative_analysis_of_proteins_1_de494646f2.jpg" alt="Bioanalytic: Quantitative analysis of proteins." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Bioanalytic_Quantitative_analysis_of_proteins_1_de494646f2.jpg 200w," sizes="100vw" width="777" height="606">

Characteristics of monoclonal antibodies (Bioanalytic)

At LabExperts, proteomic analyses are performed on a ZenoTOF 7600 high-resolution LC-MS/MS system (SCIEX). For MS/MS analyses, we use Collision induced dissociation (CID) and Electron-Activated Dissociation (EAD) fragmentation.

Antibodies are molecules made up of 4 polypeptide chains: two heavy (H - high-chain) linked to each other by a specific type of chemical bond - a disulfide bridge, and two light (L - light-chain) linked by an identical bond to the heavy chains. Monoclonal antibodies (mABs) are used in various forms - as free molecules, in the form of a drug-antibody combination, and in the form of an immunoconjugate, a drug-antibody-biopolymer combination.Their main use is in highly specific targeted therapy for cancer, cardiovascular, autoimmune and nervous system diseases. They are also used in the treatment of osteoporosis, Alzheimer's disease or transplantation. The range of applications of monoclonal antibodies is systematically expanded. The introduction of a new antibody to the market requires complex product characterization, which involves numerous proteomic analyses, which are performed by LC-MS/MS techniques, among others.<h4>LC-MS/MS</h4>The characterization of monoclonal antibodies involves many aspects, including, but not limited to, a suite of analyses related to confirmation of the structure, chain sequence and degree of modification of the antibody. Post-translational modifications (PTMs) are chemical changes that occur after the translation process. These changes generally result in an increase in the immunogenicity of the drug. For this reason, it is essential to monitor modifications as critical quality attributes (CQAs) during both the production and storage of the drug. To ensure monitoring of CQA points, the primary structure of the protein is characterized based on peptide mapping (peptide mapping) analysis. The protein under study is subjected to proteolytic digestion using enzymes such as trypsin, chymotrypsin, Lys-C, Glu-C. As a result of various proteolytic enzymes, a wide range of peptides is obtained. Ultimately, coverage of the entire sequence of the protein under study is obtained.<figure class="image"><img style="aspect-ratio:1536/947;" src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Bioanalytic_Characteristics_of_monoclonal_antibodies_3_d94fa1623b.jpg" alt="Bioanalytic: Characteristics of monoclonal antibodies." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Bioanalytic_Characteristics_of_monoclonal_antibodies_3_d94fa1623b.jpg 245w," sizes="100vw" width="1536" height="947"></figure>At LabExperts, proteomic analyses are performed on a <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?manufacturerItems=52&amp;search=ZenoTOF+7600">ZenoTOF 7600</a> high-resolution LC-MS/MS system (SCIEX). For MS/MS analyses, we use Collision induced dissociation (CID) and Electron-Activated Dissociation (EAD) fragmentation. This provides us with data for unambiguous identification of protein-building amino acids. We process the results using Sciex OS, Skyline and Spectronaut software.<blockquote>We perform:<ul><li>Whole protein analyses - Intact - to determine the exact mass and confirm the theoretical mass;</li><li>N-glycosylation profile determination (based on MS spectrum reconstruction):<ul><li>Glycosylated form;</li><li>Deglycosylated;</li><li>Glycosylated with reduced disulfide bridges;</li><li>Deglycosylated with reduced disulfide bridges.</li></ul></li><li>Peptide maps:<ul><li>Protein digestion with various proteases so that sequence coverage is close to 100%<ul><li>Analysis of PTMs:</li><li>Location of modifications</li></ul></li><li>[%] of modifications compared to the unmodified sequence</li></ul></li><li>TiPOS testing - (technical proof of similarity) - technical confirmation of the similarity of the protein synthesized by the customer (from several LOTs) with the target protein (provided as a template).</li></ul>In order to maintain the highest possible analytical standards, we perform monoclonal antibody analyses under the regime of an internally developed and implemented laboratory quality system based on GLP principles.See also our other proteomics services: <a target="_blank" rel="noopener noreferrer" href="https://labexperts.com.pl/service/identyfikacja-bialek/">Protein identification</a>, <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1294">Protein quantitative analysis</a></blockquote><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Bioanalytic_Characteristics_of_monoclonal_antibodies_2_cb93143fe6.jpg" alt="Bioanalytic: Bioanalytic Characteristics of monoclonal antibodies." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Bioanalytic_Characteristics_of_monoclonal_antibodies_2_cb93143fe6.jpg 245w," sizes="100vw" width="1536" height="908"><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Bioanalytic_Characteristics_of_monoclonal_antibodies_1_0e8ab7cdfa.jpg" alt="Bioanalytic: Bioanalytic Characteristics of monoclonal antibodies." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Bioanalytic_Characteristics_of_monoclonal_antibodies_1_0e8ab7cdfa.jpg 245w," sizes="100vw" width="1536" height="858">

Identification of the proteins (Bioanalytic)

We perform proteomic analyses on a state-of-the-art high-resolution ZenoTOF 7600 (SCIEX), in which the Zeno ion trap and EAD (Electron-Activated Dissociation) technology together make a powerful combination.

Protein identification is based on experimental measurement of peptide molecular weights and matching the amino acid sequence of the protein under study with protein sequences deposited in libraries (databases). Two approaches are most commonly used in proteomic studies: top-down and bottom-up. The top-down method involves analyzing full protein sequences. Bottom-up analyses, on the other hand, use digestion of proteins with proteolytic enzymes. The peptides obtained by digestion are analyzed using a mass spectrometer. Each analysis leads to a comparison of experimental data with data obtained by theoretical digestion of proteins described in the database.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Identification_of_the_proteins_Bioanalytic_2_6231b73f0d.jpg" alt="Bioanalytic: Identification of the proteins." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Identification_of_the_proteins_Bioanalytic_2_6231b73f0d.jpg 245w," sizes="100vw" width="1536" height="766"><h4>LC-MS/MS</h4>Tandem mass spectrometers are commonly used to determine protein sequences. They allow ion selection of pseudomolecular peptides and then fragmentation and measurement of the masses of the resulting fragments. Fragmentation using collision induced dissociation(CID ) is an invaluable tool in targeted and untargeted MS/MS analysis of small molecules. It is important to note that in proteomic analysis, data obtained using the aforementioned fragmentation method may leave gaps. For example, spectra obtained by CID fragmentation do not allow unambiguous differentiation of some amino acids: leucine from isoleucine (identical molar masses and CID fragmentation) and lysine from glutamine (very similar masses). For this reason, we perform proteomic analyses on a state-of-the-art high-resolution <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?manufacturerItems=52&amp;search=+ZenoTOF+7600">ZenoTOF 7600</a> (SCIEX), in which the Zeno ion trap and EAD (Electron-Activated Dissociation) technology together make a powerful combination. It enables the creation of unique fragments to differentiate problematic amino acids and increase the sensitivity and specificity of the analysis.<blockquote>We perform analyses of:<ul><li>samples from solution;</li><li>samples from a gel;</li><li>samples from a complex matrix (by prior arrangement);</li><li>samples from lyophilizer;</li><li>samples previously prepared by the client.</li></ul>Qualitative analysis of proteins includes:<ul><li>To confirm the presence of a protein, the sample is subjected to digestion and the result obtained from experimental data is compared with theoretical data;</li><li>Comparison of experimental data on the masses of peptides included in the analyzed protein with the protein sequences described in the database;</li><li>Analysis and identification of the protein mixture;</li><li>Identification of post translational modifications (PTMs) (we use the tools: Skyline, Spectronaut).</li><li>Reporting of the result including, but not limited to, confirmation of the coverage of the protein sequence obtained in the test sample with the theoretical protein and the list of modified peptides, location and % of modifications.</li></ul>Comparative analyses include:<ul><li>Determining the composition of mixtures;</li><li>Comparison and profiling of sample series - protein identification and trend analysis.</li></ul>Our offer also includes:<ul><li>Support in the planning and execution of studies (the scope is determined individually);</li><li>Assistance in the development of extraction methods from complex matrices - biological fluids, plant tissue, animal tissues (the scope is determined individually);</li><li>Statistical analysis using tools: MarkerView (PCA), GraphPad Prism, Trends, Sciex OS;</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1294">Quantitative analysis of proteins</a>;</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1296">Analysis and characterization of monoclonal antibodies</a>.</li></ul></blockquote><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Identification_of_the_proteins_Bioanalytic_1_97cf8c94b3.jpg" alt="Bioanalytic: Identification of the proteins." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Identification_of_the_proteins_Bioanalytic_1_97cf8c94b3.jpg 117w," sizes="100vw" width="1500" height="2000">

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