Artificial chemotaxis under electrodiffusiophoresis.

HYPOTHESIS: Through a large parameter space, electric fields can tune colloidal interactions and forces leading to diverse static and dynamical structures. So far, however, field-driven interactions have been limited to dipole-dipole and hydrodynamic contributions. Nonetheless, in this work, we propose that under the right conditions, electric fields can also induce interactions based on local chemical fields and diffusiophoretic flows. EXPERIMENTS: Herein, we present a strategy to generate and measure 3D chemical gradients under electric fields. In this approach, faradaic reactions at electrodes induce global pH gradients that drive long-range transport through electrodiffusiophoresis. Simultaneously, the electric field induces local pH gradients by driving the particle's double layer far from equilibrium. FINDINGS: As a result, while global pH gradients lead to 2D focusing away from electrodes, local pH gradients induce aggregation in the third dimension. Evidence points to a mechanism of interaction based on diffusiophoresis. Interparticle interactions display a strong dependence on surface chemistry, zeta potential and diameter of particles. Furthermore, pH gradients can be readily tuned by adjusting the voltage and frequency of the electric field. For large Péclet numbers, we observed a collective chemotactic-like collapse of particles. Remarkably, such collapse occurs without reactions at a particle's surface. By mixing particles with different sizes, we also demonstrate, through experiments and Brownian dynamics simulations, the emergence of non-reciprocal interactions, where small particles are more drawn towards large ones.

CE-MS and CE-MS/MS for the multiattribute analysis of monoclonal antibody variants at the subunit level.

The analysis of product-related substances and impurities is a critical step in the biopharmaceutical quality control of multiattribute monoclonal antibodies (mAbs), as posttranslational modifications or other variants can influence the product's biological activity. Many approaches are available for variant analysis; however, they are either variant-specific, mAb-specific, time-consuming, or require expensive equipment. Here, we present a generic capillary electrophoretic method based on a neutral-coated capillary which was coupled to mass spectrometry (MS) via the nanoCEasy interface for mAb variant analysis at the subunit level (enzymatically digested and reduced mAb). The method enabled the separation of several (i) size variants (e.g. glycosylation variants) and (ii) charge variants (e.g. c-terminal lysin clipping) as well as (iii) multiple other proteoforms (e.g. additional glycation) and (iv) incompletely reduced subunits. Separated variants were confirmed by MS/MS fragmentation even for small mass deviations like deamidation or open disulfide bridges. The system, initially developed for one mAb, was tested with nine other IgG1s to show the general applicability of the system. The presented multiattribute method enables fast and detailed characterization of mAb variants with little sample preparation and relatively simple separation equipment enabling the separation of a large set of mAb variants.

Reducing the water quenching processes using heavy water in capillary electrophoresis with fluorescence detection.

Water, ubiquitous in analytical methods, is renowned for its fluorescence quenching properties, influencing techniques like fluorescence spectrophotometry or techniques with fluorescence detection. This study explores the impact of water (H2O) substitution for heavy water (D2O) on the fluorescence behavior of anthraquinones and anthracyclines. Anthraquinones and anthracyclines play crucial roles in pharmacy, serving as essential components in various therapeutic formulations, particularly in cancer treatment and other pharmacological interventions. Capillary electrophoresis (CE) with heavy water as the background electrolyte (BGE) solvent offers superior sensitivity to the separation and detection of these analytes. Experimental results demonstrate the improved detection limits and separation efficiency of selected anthraquinones rhein (RH), aloe-emodin (AE), and anthracyclines doxorubicin (DOX), epirubicin (EPI) and daunorubicine (DAU) in heavy water-based buffers, highlighting the potential of heavy water in advancing analytical chemistry.

Smartphone-deployable and all-in-one machine vision for visual quantification analysis based on distance readout of electrophoresis titration biosensor.

As a class of point-of-care (POC) assays with visible distance readout (thermometer style), the electrophoresis titration (ET) biosensor affords high robustness, versatility, and simplicity for point-of-care quantification. However, naked-eye observation of the distance readout is unreliable in POC settings and manual processing of distance readout is time-consuming. Herein, we developed a smartphone-deployable and all-in-one machine vision for four ET biosensors (bovine serum albumin, melamine, uric acid, glutathione) to classify and quantify the samples simultaneously. To ensure accurate and rapid quantification on the smartphone, we customized the decolorization methods and edge detection operators to balance the region of interest (ROI) extraction performance and processing speed. We then established a dataset of 180 distance readout images to endow our machine vision with the ability to classify four sample types. Consequently, our machine vision demonstrated high accuracy in determining the sample type (>97.2%) and concentration (>97.3%). Moreover, expanding its applications to other targets was readily achieved by including distance readout images of other ET biosensors (e.g., hemoglobin A1c) in the dataset. Therefore, our strategy of constructing machine vision is compatible with the versatile ET biosensor technique, suggesting that the same strategy can be used for other thermometer-style POC assays.

A simple method to remove polyimide coating from fused silica capillaries for capillary electrophoresis.

A simple and cost-effective method for making capillary electrophoresis (CE) detection windows is reported. A low-cost laser engraving machine equipped with a 450 nm high-power blue laser was used to remove the polyimide coating from the fused silica capillary by laser scanning the desired section. The entire process is automated and controlled in real time by software. This method can be easily adopted by many CE users for routine practice and is scalable for high-throughput production of detection windows across multiple capillaries.

The method of capillary electrophoresis for quantitative determination of hydrophobized hyaluronic acid in its micellar forms.

Micelles based on hydrophobized hyaluronic acid (HA) are frequently used in targeted drug delivery systems. Capillary zone electrophoresis (CZE) was utilized for the quantitative determination of hydrophobized and native HA. A universal methodology was developed, suitable for the quantitative analysis of amphiphilic derivatives of hyaluronan (oleyl hyaluronan and hyaluronan conjugate with naphthalimide fluorophore) and native HA with varying molecular weights (15, 150, and 800 kDa). Furthermore, methodologies were proposed for the simultaneous quantification of a drug substance and oleyl hyaluronan in micellar forms based on the latter. The CE technique was applied for analyzing oleyl-hyaluronan-based micellar forms of two poorly soluble drug substances with oppositely charged ionic forms (loperamide and rifabutin). The examples contained in the study demonstrate a range of analytical sensitivity (LOD) for hyaluronan from 11 to 40 µg/mL and for the drug substance from 0.4 to 0.6 µg/mL. The study also showcases the accurate quantitative determination of rifabutin and loperamide in oleyl-hyaluronan-based micellar forms without the need for sample preparation. Thus, the proposed methodologies can be used to quantify native HA or its amphiphilic derivatives and simultaneously determine drug substances of various nature.

A capillary electrophoresis-based assay for carrier screening of the hotspot mutations in the CYP21A2 gene.

Molecular genetic analysis of the cytochrome P450 family 21 subfamily A member 2 (CYP21A2) gene is challenging owing to the highly homologous with its pseudogene. A reliable approach for the large-scale population screening of CYP21A2 is required. This study aimed to establish and evaluate a capillary electrophoresis-based assay for hotspot mutation carrier screening of the CYP21A2 gene. A total of 22 different variants in the CYP21A2 gene were detected by a capillary electrophoresis-based assay consisting of single nucleotide primer extension (SNaPshot) and high-throughput ligation-dependent probe amplification (HLPA) in the Chinese population, and the results were validated by alternative methods. Among the 5376 subjects, 1.51 % (81/5376) individuals were identified as CYP21A2 pathogenic variant carriers, with a carrier rate of 1/66. A total of 11 different variants were identified, of which c.293-13A/C > G (33.33 %) was the most common variant, followed by c.844C > T (19.75 %), c.518T > A (19.75 %), and Del/Con (16.05 %). There was a 100 % concordance between capillary electrophoresis and alternative method results. Furthermore, a total of 63 individuals (1.17 %, 63/5376) carried the c.955C > T (p. Q319∗) variant, among which 61 (61/63, 96.83 %) had a duplicated CYP21A2 gene and are therefore not carriers of a CYP21A2 allele. In conclusion, the capillary electrophoresis-based assay is an accurate and effective approach for genotyping the CYP21A2 gene and has the potential for the large-scale population screening of CYP21A2.

Seasonal detection of pathogens in honeybees kept in natural and laboratory conditions.

The honeybee is one of the most important pollinators in the world. The frequently observed poor health of honeybee colonies can be caused by various factors, e.g. environmental pollution, nutritional stress, and climate changes. Moreover, honeybees are constantly exposed to a wide spectrum of pathogens, such as parasites, bacteria, and viruses. We examined the occurrence of various diseases in different-aged worker honeybees from two colonies kept in natural and laboratory conditions during spring, summer, and autumn in Poland. The honeybees were examined by PCR to detect infection with selected pathogens: Nosema ceranae, N. apis, N. bombi, Acarapis woodi, trypanosomatids, and neogregarines (Mattesia or Apicystis species) and by RT-PCR to identify deformed wing virus (DWV), black queen cell virus (BQCV), and acute bee paralysis virus (ABPV). DWV and N. ceranae turned out to be the dominant pathogens. Trypanosomatids and BQCV were also found in several samples. We did not detect the presence of the other pathogens: N. apis, N. bombi, A. woodi, neogregarines, or ABPV. As shown in the present study, the dynamics and occurrence of pathogens are influenced by keeping conditions, honeybee age, and seasonality.

Greening Separation and Purification of Proteins and Peptides.

The increasing awareness of environmental issues and the transition to green analytical chemistry (GAC) have gained popularity among academia and industry in recent years. One of the principles of GAC is the reduction and replacement of toxic solvents with more sustainable and environmentally friendly ones. This review gives an overview of the advances in applying green solvents as an alternative to the traditional organic solvents for peptide and protein purification and analysis by liquid chromatography (LC) and capillary electrophoresis (CE) methods. The feasibility of using greener LC and CE methods is demonstrated through several application examples; however, there is still plenty of room for new developments to fully realize their potential and to address existing challenges. Thanks to the tunable properties of designer solvents, such as ionic liquids and deep eutectic solvents, and almost infinite possible mixtures of components for their production, it is possible that some new designer solvents could potentially surpass the traditional harmful solvents in the future. Therefore, future research should focus mainly on developing new solvent combinations and enhancing analytical instruments to be able to effectively work with green solvents.

Combining capillary electrophoresis and chemometric tools for the straightforward determination of imidazolinone herbicides in plant-based milks.

BACKGROUND: Highly polar herbicides, such as imidazolinones, are used for weed control to increase agricultural productivity and crop quality. However, their misapplication can lead to residues in ready-to-eat food with a potential health risk for consumers. Hence, the fast determination of these herbicides is necessary for timely action. In this work, an eco-friendly method based on capillary zone electrophoresis combined with chemometrics was used for the determination of imazapyr and imazamox in vegetable-based beverages such as soy and quinoa milk. RESULTS: The analytical strategy consisted of only three steps: (i) protein precipitation prior to sample injection (ii) data pre-processing to reduce the background and make corrections on electrophoretic times shift, and (iii) resolution of fully overlapped capillary electrophoresis (CE) peaks by the well-known partial least square (PLS) algorithm, which extracts quantitative information attributed to the analytes. The method was successfully applied in the concentration range between 1.00 and 100 µg L-1 with coefficient of determination of the calibration (R2 cal) and prediction (R2 pred) > 0.90, residual prediction deviation of calibration (RPDcal) and of prediction (RPDpred) > 3, and relative error of prediction (REP) > 11 in the analyzed sample matrices, in the three built methods (quinoa samples, soy samples, and joint quinoa and soy samples). CONCLUSION: The proposed methodology offers a simple and quick alternative for determining imidazolinones at trace concentrations in vegetable beverages, such as quinoa and soy milk, without complex sample preparation. The results were consistent with those obtained using more complex techniques, confirming the applicability of this method. © 2024 Society of Chemical Industry.

Enrichment of low-abundance osteopontin in bovine milk via reciprocating free-flow isoelectric focusing.

Osteopontin (OPN) in milk plays an important role in intestinal and brain development in early infancy, and great attention has been focused on OPN isolation to add extra OPN in infant formula. However, large-scale OPN isolation is limited by the low efficiency of sample pretreatment. Herein, we utilized preparative reciprocating free-flow isoelectric focusing (RFFIEF) to showcase the enrichment of low-abundance OPN in bovine milk, which contained an extremely high concentration of unwanted proteins. The reciprocating IEF format and the design of the multi-channel collector allowed us to enrich OPN in 1 L milk within 6 h. We removed 97.5% of unwanted proteins and obtained an enrichment factor of 11. Thus, our RFFIEF method can be applied to the preparative pretreatment of the large-scale milk sample and potentially improve the efficiency of downstream OPN purification.

Determination of an Anti-Parasitic Active Pharmaceutical Ingredient in Wastewater Effluents Using Capillary Zone Electrophoresis.

Ireland has a successful pharmaceutical industry with over 100 pharmaceutical manufacturing sites across the island. Although this success has many benefits, the irreversible effects emissions from pharmaceutical manufacturing can have on the environment are a major drawback. Although known pollutants are regularly monitored with limits set out by the Environmental Protection Agency, one significant pollutant has been overlooked: pharmaceutical pollution. Detecting these pollutants and ensuring they are at a safe concentration for the environment is of utmost importance. In recent years, capillary electrophoresis is being recognised as a suitable alternative to high-performance liquid chromatography due to its many benefits such as faster analysis, water-based buffers and smaller sample volumes. In this paper, a capillary zone electrophoresis (CZE) method with a preconcentration step of solid-phase extraction was developed for an anti-parasitic active pharmaceutical ingredient (API) called ZB23. The API was successfully detected in a wastewater sample in less than 10 min using the CZE parameters of 25 mM borate buffer with a pH of 10.5, 15% MeOH, 10 kV voltage, 25 mbar for 5 s injection size, an Lt of 40 cm, an Ld of 31.5 cm and a detection wavelength of 214 nm.

Stacking in electrophoresis by electroosmotic flow-assisted admicelle to solvent microextraction.

An in-line sample concentration method for capillary electrophoresis called admicelle to solvent microextraction was proposed. In this technique, analytes were trapped in the cetyltrimethylammonium bromide admicelles formed in situ on the negatively charged capillary surface. A solvent plug was then partially injected hydrodynamically to collapse the admicelles, which liberated and focused the analytes at the solvent front. Voltage was applied across the capillary, completing the stacking process. Various solvents, namely, methanol, ethanol, and acetonitrile, were investigated. The optimal solvent for solvent to admicelle microextraction was 30% acetonitrile in 24 mM sodium tetraborate (pH 9.2). Sample injection time and solvent to sample injection ratio were also optimised. For this demonstration, the optimum sample injection time and solvent to sample injection ratio were 320 s and 1:2, respectively. Using the optimum conditions, UV detection sensitivity was enhanced 132-176-fold for the model anions. The LOQ, %intra-/inter-day (n = 6/n = 12, 2 days) repeatability, and linearity (R2) of admicelle to solvent microextraction were 0.08-2 µg/mL, 1.9-3.9%, 2.8-4.9%, and 0.992, respectively. Admicelle to solvent microextraction was applied to the analysis of various fortified water samples, with good repeatability (%RSD = 0.5-3.6%), and no matrix interferences.

CE-UV method for the determination of catecholamine metabolites from baby pee-covered diapers.

A method has been developed for the analysis of vanillylmandelic acid, homovanillic acid, and 5-hydroxyindoleacetic acid from baby urine as biomarkers of neuroblastoma in infants. Disposable diapers were employed as sampling devices in order to guarantee a low invasiveness during this step. The proposed method consists on a simple extraction step with water from the used diaper followed by the measurement using capillary electrophoresis with UV detection. The Box-Behnken design (BBD) was utilized to optimize the process of extracting catecholamine metabolites from the examined samples. The variables of the sample preparation step were optimized and the method was validated obtaining limits of quantification of 1.65 µg mL-1, good intraday and inter-day precision with RSDs under 15 %. Finally the method was applied to real samples collected from the Department of Neonatology, University Clinical Centre (Gdansk, Poland). The greenness of the proposed method was also evaluated with different tools (i.e., AGREEPrep and GAPI) with satisfactory results, which allow to state that the method can be considered green. Moreover, its practicality was evaluated by application of BAGI tool, proving to be a practical and economical method to be applied in routine laboratories for determination of catecholamine metabolites in urine-type samples.

Clindamycin phosphate-based deep eutectic solvent as a chiral selector for enantioseparation of amino alcohol drugs in nonaqueous capillary electrophoresis.

Clindamycin phosphate (CP) exhibits good enantioselectivity for many basic drugs, but its separation effect for most amino alcohol drugs is not satisfactory. In this work, a deep eutectic solvent (DES) chiral selector based on CP was prepared for the first time and utilized as a single chiral selector in nonaqueous capillary electrophoresis (NACE) to separate twelve amino alcohol drugs. Compared with unmodified CP, the separations of model drugs in the DES chiral selector system were significantly improved. Most amino alcohol drugs could be completely separated, and the peak shapes were good. In addition, we used infrared spectroscopy and nuclear magnetic resonance method to study the specific separation mechanism and explored the reasons why DES chiral selector has better enantioselectivity. This work is the first to directly modify antibiotic chiral selector into DES, indicating a direction for us to develop novel chiral recognition materials.

Health assessment of nesting loggerhead sea turtles (Caretta caretta) in one of their largest rookeries (central eastern Florida coast, USA).

Reproduction is a physiologically demanding process for sea turtles. Health indicators, including morphometric indices and blood analytes, provide insight into overall health, physiology and organ function for breeding sea turtles as a way to assess population-level effects. The Archie Carr National Wildlife Refuge (ACNWR) on Florida's central eastern coast is critical nesting habitat for loggerhead sea turtles (Caretta caretta), but health variables from this location have not been documented. Objectives of the study were to (1) assess morphometrics and blood analyte data (including haematology, plasma biochemistry, protein electrophoresis, ß-hydroxybutyrate, trace nutrients, vitamins and fatty acid profiles) from loggerheads nesting on or near the beaches of the ACNWR, (2) investigate correlations of body condition index (BCI) with blood analytes and (3) analyse temporal trends in morphometric and blood analyte data throughout the nesting season. Morphometric and/or blood analyte data are reported for 57 nesting loggerheads encountered between 2016 and 2019. Plasma copper and iron positively correlated with BCI. Mass tended to decline across nesting season, whereas BCI did not. Many blood analytes significantly increased or decreased across nesting season, reflecting the catabolic state and haemodynamic variations of nesting turtles. Twenty-three of 34 fatty acids declined across nesting season, which demonstrates the physiological demands of nesting turtles for vitellogenesis and reproductive activities, thus suggesting potential utility of fatty acids for the assessment of foraging status and phases of reproduction. The findings herein are relevant for future spatiotemporal and interspecies comparisons, investigating stressor effects and understanding the physiological demands in nesting sea turtles. This information provides comparative data for individual animals in rescue or managed care settings and for assessment of conservation strategies.

Stage-Specific Plasma Metabolomic Profiles in Colorectal Cancer.

Background/Objectives: The objective of this study was to investigate the metabolomic profiles of patients with colorectal cancer (CRC) across various stages of the disease. Methods: The plasma samples were obtained from 255 subjects, including patients with CRC in stages I-IV, polyps, and controls. We employed capillary electrophoresis time-of-flight mass spectrometry and liquid chromatography triple quadrupole mass spectrometry to analyze hydrophilic metabolites comprehensively. The data were randomly divided into two groups, and consistent differences observed in both groups were analyzed. Results: Acetylated polyamines, such as N1-acetylspermine and N1, N12-diacetylspermine, consistently showed elevated concentrations in stage IV compared to stages I-III. Non-acetylated polyamines, including spermine and spermidine, exhibited increasing trends from polyp to stage IV. Other metabolites, such as histidine and o-acetylcarnitine, showed decreasing trends across stages. While acetylated polyamines have been reported as CRC detection markers, our findings suggest that they also possess diagnostic potential for distinguishing stage IV from other stages. Conclusions: This study showed stage-specific changes in metabolic profiles, including polyamines, of colorectal cancer.

Interactions of electrophoretically silent hemoglobin Hekinan II [HBA1:c.84G>T] with various forms of α-thalassemias and other hemoglobinopathies: novel insights into the molecular and hematological characteristics and genetic origins.

To determine the molecular basis, genotype - phenotype relationship, and genetic origin of Hemoglobin (Hb) Hekinan associated with several forms of α-thalassemia and other hemoglobinopathies for a better understanding of its diverse clinical phenotypes. Seventeen participants with suspected abnormal Hb were studied. Hb analysis was performed using high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE). Mutational and α-haplotypic and structural analyses were conducted, and the effects of mutations on globin-chain stability were determined. All participants harbored Hb Hekinan II (HBA1:c.84 G>T) co-inherited with another α-globin gene anomaly. Three novel genotypes, (ααHekinan/αCSα), (ααHekinan/αCSα,ßA/ßE), and (ααHekinan/αCSα,ßE/ßE), were characterized. Despite being co-inherited with both α- and ß-Hb variants Hb Hekinan II led to minimal changes in erythrocyte parameters, suggesting a non-pathological nature. HPLC but not CE revealed a distinct small shoulder-like Hb pattern. Thai Hb Hekinan II was strongly associated with haplotype [+ - S + - - -] and the possibility of four different haplotypes, while two Burmese Hb Hekinan II were associated with haplotypes [± - S + - + -] and [± - S + - - -]. The novel genotypes identified provide a fresh perspective on Hb Hekinan II diversity. HPLC has superior identification capabilities for samples of Hb Hekinan II co-inherited with α-thalassemia. Thai and Burmese Hb Hekinan II have diverse origins.

Pilot Study for Deciphering Post-Translational Modifications and Proteoforms of Tau Protein by Capillary Electrophoresis-Mass Spectrometry.

Abnormal accumulation of tau protein in the brain is one pathological hallmark of Alzheimer's disease (AD). Many tau protein post-translational modifications (PTMs) are associated with the development of AD, such as phosphorylation, acetylation, and methylation. Therefore, a complete picture of the PTM landscape of tau is critical for understanding the molecular mechanisms of AD progression. Here, we offered a pilot study of combining two complementary analytical techniques, capillary zone electrophoresis (CZE)-tandem mass spectrometry (MS/MS) and reversed-phase liquid chromatography (RPLC)-MS/MS, for bottom-up proteomics of recombinant human tau-0N3R. We identified 50 phosphorylation sites of tau-0N3R in total, which is about 25% higher than that from RPLC-MS/MS alone. CZE-MS/MS provided more PTM sites (i.e., phosphorylation) and modified peptides of tau-0N3R than RPLC-MS/MS, and its predicted electrophoretic mobility helped improve the confidence of the identified modified peptides. We developed a highly efficient capillary isoelectric focusing (cIEF)-MS technique to offer a bird's-eye view of tau-0N3R proteoforms, with 11 putative tau-0N3R proteoforms carrying up to nine phosphorylation sites and lower pI values from more phosphorylated proteoforms detected. Interestingly, under native-like cIEF-MS conditions, we observed three putative tau-0N3R dimers carrying phosphate groups. The findings demonstrate that CE-MS is a valuable analytical technique for the characterization of tau PTMs, proteoforms, and even oligomerization.

Serum chemistry, thyroxine concentration, and blood cell morphology of wild, young Blackchin Guitarfish Glaucostegus cemiculus sampled on the eastern Mediterranean coastline.

OBJECTIVE: The Blackchin Guitarfish Glaucostegus cemiculus is endemic to the Mediterranean Sea and is critically endangered, but relevant routine laboratory data are unavailable. Our objectives were to determine the packed cell volume (PCV), comprehensive serum chemistry analytes, and serum total thyroxine (sTT4) concentration; compare serum albumin and serum globulin concentrations as measured by two different methods; and describe the blood cell morphology of healthy, free-ranging Blackchin Guitarfish. METHODS: Wild Blackchin Guitarfish were captured using a seine net. Blood samples for serum chemistry and hematological analyses were obtained and measured using routine laboratory methods. The fish were tagged and released. RESULT: This study included 43 Blackchin Guitarfish (17 males and 26 females) that were younger than 6 months as estimated based on total length and body weight. The median PCV (n = 23) was 22% (minimum-maximum [min-max] = 15-25%). Median sTT4 (n = 10) measured by chemiluminescence immunoassay was 7.86 nmol/L (min-max = 7.52-9.57 nmol/L). The study included a comprehensive, 25-analyte serum chemistry analysis (e.g., serum iron and unbound and total iron-binding capacity) and a morphological description of all blood cells. Serum electrophoresis (SEP; n = 13) yielded a consistent serum albumin-migrating protein fraction and four globulin fractions. Serum electrophoretograms corroborating these results are presented. CONCLUSION: In Blackchin Guitarfish, the serum albumin-migrating fraction measured by SEP combined with serum total protein concentration yields a much higher albumin concentration compared to that measured by bromocresol green spectrophotometry. The true identity of this albumin-migrating fraction remains to be identified. The analytes' calculated 2.5-97.5% interpercentile intervals should be considered as reference intervals applying to Blackchin Guitarfish of similar age but should be applied cautiously to adult fish.