PLASMA PROTEIN ELECTROPHORESIS IN THE WHITE STORK (CICONIA CICONIA): AGREEMENT BETWEEN AGAROSE GEL VERSUS CAPILLARY ZONE METHODS AND DEVELOPMENT OF REFERENCE INTERVALS.

The white stork (Ciconia ciconia) is a ciconiiform species widely represented in zoological institutions. Plasma protein electrophoresis is widely used in avian patients for assessment of inflammatory conditions, but reference intervals for this testing modality are lacking for the white stork. The two main electrophoretic methods are agarose gel electrophoresis (AGE) and capillary zone electrophoresis (CZE). This study assessed fresh plasma samples of healthy adult white storks (n = 30). Statistical analyses were performed to evaluate agreement between AGE and CZE. Typical electrophoretic fractions were obtained from both methods (prealbumin, albumin, α1, α2, ß, γ1, and γ2). The AGE and CZE methods were not equivalent for determining major electrophoretic fractions (except ß-globulins) and albumin:globulin ratio on plasma samples. An additional prealbumin fraction was seen with CZE. Reference intervals were established for each method as the smallest n group was 27 individuals for a given value; most values had normal distribution, and robust or parametric methods were used on the data.

Method comparison for serum protein electrophoretic M-protein quantification: Agarose gel electrophoresis and capillary zone electrophoresis in canine and feline sera.

BACKGROUND: Densitometric quantification of myeloma paraproteins (M-proteins) is used to monitor secretory myeloma related disorders in humans and dogs. The previous work in dogs used agarose gel electrophoresis (AGE) but did not establish if other methods of serum protein electrophoresis, such as capillary zone electrophoresis (CZE), were comparable. OBJECTIVES: We aimed to determine if the densitometric quantification of M-proteins using CZE would yield results comparable to AGE methods. METHODS: Fifty-one serum samples from 22 dogs and 18 cats with confirmed monoclonal gammopathies and previously performed AGE were evaluated using CZE on a Sebia Minicap system. Samples were run in duplicate, and their M-proteins were densitometrically measured using the corrected perpendicular drop method previously described. Human-based quality control samples were used to determine the inter-run coefficient of variation (CV). Patient samples were used to calculate the intra-run CV. Method comparison was performed using simple linear regression, Passing-Bablok regression, and Bland-Altman analyses, and Medx evaluations. RESULTS: Inter-run and intra-run CVs for CZE were 3.71%-7.65% and 2.89%-4.74%, respectively. Simple linear regression demonstrated an excellent correlation (r > 0.98). Passing-Bablok regression was compatible with the presence of proportional bias in the entire population, and Bland-Altman plots revealed a proportional bias in the feline cases. The Medx evaluation suggested that the two methods did not perform similarly in clinical samples with poor performance at a decision limit of 0.5 gm/dL. CONCLUSIONS: Capillary zone electrophoresis is an acceptable method for M-protein densitometric quantification in canine and feline sera but cannot be used interchangeably with AGE-based evaluations.

Protein Electrophoresis of Serum and Heparinized Plasma in the Common Mynah (Acridotheres tristis).

Although serum protein electrophoresis is a diagnostic tool available through many veterinary laboratories, there currently are no reference intervals for protein fractions in healthy common mynahs (Acridotheres tristis). Therefore, electrophoretic patterns of proteins in serum and heparinized plasma of the common mynah were evaluated. Blood specimens were collected from 55 healthy adult common mynahs of unknown age (26 males and 29 females). The serum total protein and protein fractions were measured using the biuret method followed by cellulose acetate electrophoresis (CAE). The serum level of albumin was compared with bromocresol green (BCG) dye-binding and CAE methods. Four protein fractions, including albumin and α, ß, and γ globulins, were recorded in the electrophoretogram of serum specimens. Sex appeared to have no significant effect on the measured parameters. The serum BCG albumin fraction was significantly higher than the CAE albumin fraction (P = .01). Also, the comparison of total protein and protein fractions in serum and plasma specimens of 25 of the 55 birds sampled showed that total protein (Cohen index d = 0.66, P = .03), gamma globulin (d = 1.13, P = .00), and total globulin (d = 0.67, P = .00) in plasma samples were significantly higher than those in serum samples. The results of this study provide the specific reference intervals for total protein and protein fractions in common mynahs, which are essential for proper interpretation of laboratory results and also revealed that the albumin measurement by the BCG method yields unreliable results in common mynahs.

COMPARISON OF PROTEIN ELECTROPHORESIS AND BIOCHEMICAL ANALYSIS FOR THE QUANTIFICATION OF PLASMA ALBUMIN IN HEALTHY BEARDED DRAGONS (POGONA VITTICEPS).

While electrophoresis is considered the standard method for evaluation of protein concentrations as a result of its direct measurement, albumin is often quantified with biochemical assays. Many laboratory-based chemistry analyzers and clinic-based point-of-care analyzers use the dye bromocresol green (BCG) for the quantitation of albumin. Several studies have shown that albumin concentrations obtained by the standard (BCG) dye-binding method are significantly different from those obtained by protein electrophoresis in avian species and chelonia. The goal of this study was to compare plasma albumin concentrations obtained by the BCG method with those derived from electrophoresis in bearded dragons (Pogona vitticeps). Thirty-six heparinized plasma samples were obtained from 13 clinically healthy male bearded dragons. Albumin was quantified by protein electrophoresis and by the BCG dye-binding method. The two methods were significantly different (P < 0.0001, paired t-test; P < 0.0001, Wilcoxon signed-rank test), with the BCG measurement always equal to or higher than the electrophoretic result. The measurements from both methods were significantly correlated (r = 0.8634, P < 0.0001), but concordance between the two techniques was poor. The Bland-Altman plot appeared to show a greater difference between the two measurements with lower albumin values and lesser difference with higher values. These results indicate that bearded dragon plasma albumin concentration measurements obtained by the BCG dye-binding method are unreliable when compared to those obtained with electrophoresis, suggesting that albumin should be measured by protein electrophoresis for health assessment in bearded dragons.

Diagnostic performance of routine electrophoresis and immunofixation for the detection of immunoglobulin paraproteins (M-Proteins) in dogs with multiple myeloma and related disorders: Part 2-Toward improved diagnostic performance.

BACKGROUND: The diagnostic performance of routine electrophoresis (agarose gel electrophoresis [AGE] and capillary zone electrophoresis [CZE]) and species-specific immunofixation (IF) for the detection of immunoglobulin paraproteins (M-proteins) and diagnosis of secretory myeloma-related disorders (sMRD) can be improved. Available canine IF targets were IgG-FC, IgA, IgM, light chain (LC), IgG4, and free LC (fLC) antibodies. OBJECTIVE: We aimed to review specific features associated with the presence of M-proteins in canine serum samples and the common features causing inaccurate reporting of M-proteins to improve the diagnostic performance of routine electrophoresis and IF for the detection of M-proteins. METHODS: Features found in AGE, CZE, routine IF, IgG4 IF, and fLC IF of 100 canine serum samples from Part 1 of this study were evaluated by simple and multivariate logistic regression to identify factors associated with the presence of M-proteins. Cases falsely called negative or positive for M-proteins were reviewed to identify the common features that could be used to increase the diagnostic performance of SPE and IF for M-protein detection. RESULTS: The presence of hypogammaglobulinemia or any peak taller than albumin was associated with an M-protein. Total protein concentrations, globulin concentrations, or peaks wider than albumin were not associated with an M-protein. Free LC sMRD cases were not diagnosed by SPE and routine IF. Cases with infectious and inflammatory etiologies had a restricted polyclonal gammopathy with multiple γ-globulin restrictions resulting in some false-positive results. SPE combined with all available IF results and the specific features identified in this study had an estimated sensitivity of 95.1% and specificity of 81.4%. CONCLUSIONS: The identified criteria of this study increase the diagnostic performance of the electrophoretic evaluation for M-proteins.

Differences in serum protein electrophoretic pattern in dogs naturally infected with Babesia gibsoni and Babesia canis.

Canine babesiosis may cause several hematological and biochemical changes, but only limited studies are available regarding the possible differences of changes in animals infected by different Babesia parasites. The study focused on the evaluation of the differences in serum protein electrophoretic pattern between dogs naturally infected with B. gibsoni (17 dogs) and B. canis (40 dogs). The mean values of total proteins, ß1-, ß2- and γ-globulins were in dogs infected with B. gibsoni significantly higher (P < 0.05 and P < 0.001) than in dogs infected with B. canis. The relative concentrations of albumin, α1-, α2-globulins and the A/G ratios were in the B. gibsoni infected dogs significantly lower (P < 0.001), no significant differences were found in the relative concentrations of ß1- and ß2-globulins. Significant differences were found in most of the evaluated parameters when comparing the results in relation to the form of B. canis infection to B. gibsoni infection. Hematological indices showed significant differences between dogs infected with B. gibsoni and the complicated form of B. canis infection. In conclusion, the obtained results suggest differences in the changes of serum protein electrophoretic pattern between dogs infected with both Babesia species and thus, in the response to the infection caused by various Babesia parasites.

ESTABLISHMENT OF ACUTE-PHASE PROTEIN AND SERUM PROTEIN ELECTROPHORESIS PRELIMINARY REFERENCE VALUES FOR PRONGHORN (ANTILOCAPRA AMERICANA).

Pronghorn (Antilocapra americana) are native to western North America and are found in 24 Association of Zoos and Aquariums (AZA)-accredited institutions. Acute-phase proteins (APP) are a broad class of proteins that are stimulated in response to inflammation and have been shown to be a sensitive measure of inflammation in equids and ruminants. In this study, blood samples from clinically normal free-ranging and captive populations of pronghorn were analyzed using assays for protein electrophoresis (EPH) and APP, including serum amyloid A (SAA) and haptoglobin (HP), to develop preliminary ranges to gauge potential differences between these populations. Additional samples were taken from clinically abnormal captive pronghorn with facial abscesses. By EPH measurements, albumin: globulin ratio mean and SE were significantly different (P <0.05) with 1.02 (0.08) for captive populations and 1.91 (0.05) for free-ranging populations. Total protein mean and SE were significantly different (P <0.05) for captive and free-ranging populations, respectively 5.6 (0.3) g/dl and 6.9 (0.1) g/dl. Mean and SD of SAA for captive pronghorn were 1.4 (3.2) mg/L, and were significantly different from the free-ranging population, which was below the limits of detection for (P <0.05). There was no difference in HP levels between these groups. In a case study of a pronghorn with facial abscesses, elevated levels of HP, but not SAA, suggested that HP maybe useful in certain disease states. Future studies should explore the use of these biomarkers as tools to monitor general health, prognosis, and subclinical disease.

COMPARISON OF AGAROSE GEL AND CAPILLARY ZONE ELECTROPHORESIS METHODS USING PLASMA FROM GREEN TURTLES (CHELONIA MYDAS).

Agarose gel electrophoresis (AGE) has been widely implemented throughout veterinary medicine and for analysis of plasma proteins of avian and reptile species. Capillary zone electrophoresis (CZE) is becoming a standard method in human clinical pathology laboratories but has not widely been used for the analysis of animal samples. The objective of the present study was to compare protein fractions derived from AGE and CZE methods using plasma from the green turtle (Chelonia mydas). Plasma samples were analyzed by AGE and CZE per manufacturer guidelines. The methods were assessed by CV analysis, Spearman's correlation, Passing-Bablok regression, and Bland Altman plots. CZE consistently resolved more fractions than AGE with three fractions observed in the prealbumin migrating region versus one for AGE and two fractions in the γ globulin region versus one for AGE. Compared with AGE, CZE showed a lower CV in intra-assay tests (1.0-4.9% vs 2.0-28.3%) and a lower or overlapping CV in interassay tests (1.0-10.6 vs 2.3-22.0). The prealbumin, α2 globulin, and ß globulin fractions correlated the least between the methods (for all three fractions: rs ≤ 0.28, P > 0.21). Moderate, significant correlations between AGE and CZE methods were observed for albumin (rs = 0.78, P < 0.0001) and γ globulins (rs = 0.78, P < 0.0001). CZE has a higher precision and ease of use over AGE and offers the opportunity to resolve additional protein fractions. This will necessitate the development of new conventions in placement of fraction delimits, definition of species-specific reference intervals, and evaluation of clinical utility in abnormal turtles.

Serum Protein Analysis of Nurse Sharks.

Serum protein electrophoresis (EPH) is used to assess relative concentrations of blood proteins in clinical and biological studies. Serum EPH fractions have been determined for elasmobranchs using mammalian albumin, alpha 1-, alpha 2-, beta-, and gamma-globulin fractions, and have been deemed fractions 1 through 5, respectively. However, serum EPH fraction concentration reference intervals (RIs) have not been widely established for different elasmobranch species. In this study, RIs for fractions 1 through 5 were determined from 45 wild-caught Nurse Sharks Ginglymostoma cirratum (27 females and 23 males) in South Florida. Serum samples were isolated from whole blood following caudal venipuncture. Body condition was also measured in the field to assess the relative health of the individuals sampled. There was no relationship between body condition and serum EPH fraction concentrations. In addition, there was no difference in body condition or serum EPH fraction concentrations between females and males. Total solids and total protein values were significantly different (P < 0.001). Nurse Shark serum EPH fraction 1 was found within the mammalian albumin migrating band distance and was negligible. Fraction 2 showed no peak in the mammalian alpha 1-globulin range. A thin, medium peak in the mammalian alpha 2-globulin range represented fraction 3. In the mammalian beta-globulin range, fraction 4 consisted of the majority of protein observed. It was represented by a smooth, broad peak. A short, medium broad peak in the mammalian gamma-globulin range represented fraction 5. The Nurse Shark serum EPH fraction RIs provided in this study may be utilized to clinically evaluate the health of Nurse Sharks in captivity and in the wild, and to compare the health of their populations around the world experiencing various anthropogenic stressors and other environmental impacts.

PLASMA BIOCHEMISTRY AND PROTEIN ELECTROPHORESIS REFERENCE INTERVALS OF THE COMMON LOON (GAVIA IMMER).

There are no published plasma biochemistry reference intervals for any species within the order Gaviiformes, which includes the common loon (Gavia immer). Because of their unique classification and lack of close taxonomic relatives, species-specific values for clinical data in loons are needed. This study determined reference intervals for plasma biochemical values in adult common loons, and reference intervals for protein electrophoresis values in both adult and juvenile common loons. Healthy, wild adult (n = 148, age >3 yr) and juvenile (n = 31, age 4-12 wk) common loons were sampled on freshwater summer breeding territories at study sites across North America. Plasma biochemical analytes included glucose (Glu), total calcium, phosphorus, sodium, potassium, chloride, blood urea nitrogen (BUN), uric acid, cholesterol, triglycerides, creatine kinase, γ-glutamyl transferase, alkaline phosphatase (ALP), lactate dehydrogenase, aspartate aminotransferase, amylase, and bile acids. Protein electrophoresis data included albumin to globulin ratio (A: G), prealbumin, albumin, α1-globulin, α2-globulin, ß-globulin, and γ-globulin. Adult females had significantly higher Glu, ALP, and BUN than adult males. Juvenile loons had higher ß-globulins than adults, whereas adults had higher α1-globulins. Establishment of complete reference intervals will improve clinical assessment of captive loons, and allow researchers to better understand the health of wild loons in response to the multiple environmental stressors faced by these species.

SERUM ACUTE-PHASE PROTEINS IN BOTTLENOSE DOLPHINS (TURSIOPS TRUNCATUS) AND CORRELATION WITH COMMONLY UTILIZED INFLAMMATORY INDICES.

Acute-phase proteins (APP) are the foundation to the innate immune response and valuable biomarkers that increase with inflammation, infection, neoplasia, stress, and trauma.2,4,16 Little is known about the acute-phase response in cetaceans and if these proteins can be used for health monitoring in individuals and free-ranging populations. The purpose of this study was to characterize serum concentrations of haptoglobin (Hp) and serum amyloid A (SAA), as well as electrophoretic profiles of common bottlenose dolphins (Tursiops truncatus) in free-ranging (n = 33) and professional care (n = 27) settings. Results were correlated to commonly utilized inflammatory indices including erythrocyte sedimentation rate (ESR), fibrinogen, total white blood cell count (WBC), and absolute neutrophil count. SAA levels, measured with a dolphin-specific enzyme linked immunosorbent assay (ELISA), were significantly higher (P = 0.05) in free-ranging dolphins (mean = 4.26; SE = 1.12) when compared with those under professional care (mean = 1.82; SE = 0.45). For dolphins under professional care, a statistically significant correlation was identified between ESR and Hp (P < 0.001; r = 0.69), ESR and SAA (P < 0.001; r = 0.67), fibrinogen and Hp (P = 0.001; r = 0.58), and fibrinogen and SAA (P = 0.002; r = 0.56). In addition, there was a significant correlation between WBC and SAA (P = 0.01; r = 0.38) and absolute neutrophil count and SAA (P = 0.04; r = 0.32). There were no significant correlations between study variables observed in free-ranging dolphins. The variable correlation of APPs with commonly utilized inflammatory indices demonstrates that these proteins are independent measures of inflammation with unique sensitivity, specificity, and timeline of expression. The results of this study contribute to improved health monitoring of dolphins and have the potential to assist in identification of compromised health.

Validation and method comparison of the use of densitometry to quantify monoclonal proteins in canine sera.

BACKGROUND: Densitometric quantitation using serum protein electrophoresis (SPE) is used to monitor monoclonal proteins (M-proteins) in human patients but has not been validated in the dog. Serum globulin concentrations, species-specific radial immunodiffusion (RID), and ELISAs are currently used in veterinary medicine. OBJECTIVE: We aimed to compare four methods that quantify M-proteins using densitometry and biuret protein (dM-protein) measurements. We also validated the best performing method and compared it with the RID and ELISA methods for measuring canine serum M-protein. METHOD: Serum from six normal dogs and 83 serum samples from 46 dogs with confirmed monoclonal gammopathies were used. A spike and recovery experiment with purified monoclonal IgG and IgM, inter-run and intra-run variability, linearity under dilution, and lower limit of detection were performed. Results of commercial canine RID and ELISA kits for total class-specific immunoglobulin were compared with dM-proteins. RESULTS: The corrected perpendicular drop gating method had <20% error for IgG/γ-globulin and IgM/ß-globulin M-protein quantifications. Linearity (r > .99), intra-run CV (1.1%-2.3%), and inter-run CVs (2.0%-3.5%) were acceptable. Correlation between the RID and densitometry results ranged from r = .25 to r = .88, depending on the class. The RID result was greater than that of the biuret total protein in 26/63 (41%) IgA cases. A panel of IgG, IgA, and IgM RIDs failed to correctly identify an IgM paraproteinemia in 6/6 (100%) cases. Densitometry was not comparable with any other tested method. CONCLUSION: Densitometric quantitation is a valid technique for measuring M-proteins in the ß- and γ-globulin regions. Immunotyping via RID using the tested kit does not appear to detect IgM. Densitometry is recommended for measuring M-proteins in canine patients.

Plasma protein fractions in free-living white-tailed eagle (Haliaeetus albicilla) nestlings from Norway.

BACKGROUND: Capillary electrophoresis of plasma proteins has shown great potential as a complementary diagnostic tool for avian species. However, reference intervals for plasma proteins are sparse or lacking for several free-living avian species. The current study reports electrophoretic patterns and concentrations of plasma proteins determined for 70 free-living white-tailed eagle (Haliaeetus albicilla) nestlings from two locations in Norway (Steigen and Smøla) in order to establish reference values for this subpopulation using capillary electrophoresis. The nestlings were between 44 and 87 days of age, and the plasma protein concentrations were investigated for age, sex, year (2015 and 2016) and location differences. To our knowledge, this is the first report of reference intervals of plasma proteins analysed by capillary electrophoresis in free-living white-tailed eagle nestlings. RESULTS: The plasma protein concentrations (% of total protein, mean ± SE) were determined for prealbumin (13.7%, 4.34 ± 0.15 g/L), albumin (46.7%, 14.81 ± 0.24 g/L), α1-globulin (2.4%, 0.74 ± 0.03 g/L), α2-globulin (11.7%, 3.72 ± 0.06 g/L), ß-globulin (15.9%, 5.06 ± 0.08 g/L) and γ-globulin (9.6%, 3.05 ± 0.09 g/L). Significant differences were found between the two locations for prealbumin, α2- and γ-globulins. No significant differences were found between the two sampling years or sexes, and no effect of age was found for any of the plasma proteins. However, prealbumin levels were several folds higher than previously reported from adults of closely related birds of prey species. There were no other studies on capillary electrophoresis of nestling plasma available for comparison. CONCLUSION: Significant differences were found between sampling locations for prealbumin, α2- and γ-globulins, which may indicate differences in inflammatory or infectious status between nestlings at the two locations. Sampling year, sex or age had no significant effect on the plasma protein concentrations. These results provide novel data on plasma protein concentrations by capillary electrophoresis and may be useful for evaluation of health status in free-living white-tailed eagle nestlings.

Relação do proteinograma sérico e as fases folicular e luteal do ciclo estral em éguas / Relation of the serum proteins and the follicular and luteal phasesof the estrous cycle in mares

O objetivo deste estudo foi obter o perfil eletroforético das proteínas séricas em éguas cíclicas e verificar as diferenças entre as fases folicular e luteal do ciclo estral nesta espécie. Foram utilizadas 18 éguas, totalizando 36 amostras de soro, sendo duas de cada égua. As amostras foram colhidas no estro e no diestro. As proteínas séricas totais foram obtidas pelo método do Biureto, a partir da utilização de Kits comerciais (LABTEST®) e, as diferentes subfrações proteicas, por eletroforese em gel de poliacrilamida (SDS-PAGE). O eletroforetograma das proteínas séricas colocou em evidência a presença de 17 a 25 frações proteicas, cujos pesos moleculares variaram de 22 a 254 kDa. Identificaram-se duas proteínas ainda não nomeadas oficialmente, de massas moleculares (MM) 23 kDa e 144 kDa. Os valores médios ± SEM obtidos para cada variável no estro e no diestro, respectivamente, foram: proteínas totais (g/dL) 7,11 ± 0,07 e 7,36 ± 0,07; albumina (mg/dL) 4790,83 ± 69,10 e 5027,19 ± 69,10; α1 glicoproteína ácida (mg/dL) 4,90 ± 0,31 e 4,93 ± 0,31; ceruloplasmina (mg/dL) 15,28 ± 1,31 e 10,65 ± 1,31; haptoglobina (mg/dL) 22,70 ± 1,16 e 27,06 ± 1,16; transferrina (mg/dL) 329,00 ± 9,78 e 350,16 ± 9,78; IgA (mg/dL) 119,91 ± 6,30 e 107,03 ± 6,30; IgG (mg/dL) 1525,07 ± 40,18 e 1517,25 ± 40,18; MM 23 (mg/dL) 204,44 ± 8,61 e 219,79 ± 8,61; MM 144 (mg/dL) 22,13 ± 0,55 e 21,49 ± 0,55. Não houve diferença significativa das proteínas totais e suas frações do estro para o diestro. Conclui-se que as modificações hormonais durante as fases do ciclo estral da égua não interferem no proteinograma sérico.

This study aimed to obtain the electrophoretic profile of serum proteins in cyclic mares and to verify the differences between the follicular and luteal phases of the estrous cycle in this species. Eighteen mares were used, totaling 36 serum samples, two of each mare. Samples were collected both in estrus and in diestrus. Total serum proteins were obtained by the Biureto method, by using commercial kits (LABTEST®), while the different protein subfractions by polyacrylamide gel electrophoresis (SDS-PAGE). The electroforetogram of serum proteins evidenced the presence of 17 to 25 protein fractions, whose molecular weights ranged from 22 to 254 kDa. Two proteins that were not yet officially named were identified, of molecular weights (MW) of 23 kDa and 144 kDa. The mean values (± SEM) obtained for each variable in estrus and diestrus were, respectively: total proteins (g/dL) 7.11 ± 0.07 and 7.36 ± 0.07; albumin (mg/dL) 4790.83 ± 69.10 and 5027.19 ± 69.10; α1 acid glycoprotein (mg/dL) 4.90 ± 0.31 and 4.93 ± 0.31; ceruloplasmine (mg/dL) 15.28 ± 1.31 and 10.65 ± 1.31; haptoglobine (mg/dL) 22.70 ± 1.16 and 27.06 ± 1.16; transferrin (mg/ dL) 329.00 ± 9.78 and 350.16 ± 9.78; IgA (mg/dL) 119.91 ± 6.30 and 107.03 ± 6.30; IgG (mg/dL) 1525.07 ± 40.18 and 1517.25 ± 40.18; MW 23 (mg/dL) 204.44 ± 8.61 and 219.79 ± 8.61; MW 144 (mg/dL) 22.13 ± 0.55 and 21.49 ± 0.55. No significant difference was verified in total proteins and its fractions in estrus and diestrus. The hormonal changes during the specific stages of the estrous cycle of the mare do not interfere with the serum proteinogram.

Serum proteinogram of the Campeiro horse / Proteinograma sérico de equinos da raça Campeiro

The aim of this study to measure the fractions of the total serum proteins of the Campeiro horse and identify the influences of biological variants. Blood samples were taken in 138 horses of the breed Campeiro for measuring the concentration of total serum protein by the biuret method. Serum concentrations of protein fractions were measured by electrophoresis using agarose gel. Groups were formed according to age, sex and reproductive condition. The average values of serum fractions: albumin (2.85±0.36g/dl), alpha 1 (0.28±0.11g/dl), alpha 2 (0.26±0.08g/dL) beta 1 (0.57±0.15g/dl), beta 2 (0.89±0.28g/dL), gamaglobulinas (1.86±0.34g/dL), albumin/globulin ratio (0.75±0.18) and 2.5% percentile and 97.5% had slight differences in relation to the reference interval proposed for the species. They observed higher values of alpha 1 and 2 globulins in the group from that had six to eight years old and gammaglobulins in group above 13 years old. Serum protein concentrations were similar in horses and mares and between non-pregnant and pregnant. Sex and pregnancy status did not affect serum proteinogram. Alpha and gammaglobulins have higher values as the age increases. Serum proteinogram of Campeiro horses shows variations that have to be considered in the interpretation of laboratory tests.(AU)

Este trabalho tem por objetivo mensurar as frações das proteínas totais séricas de equinos Campeiros e identificar as influências de variantes biológicas. Foram colhidas amostras de sangue de 138 equinos, machos e fêmeas da raça Campeiro. A determinação da concentração de proteínas totais séricas foi realizada pelo método de biureto. As concentrações séricas das frações proteicas foram determinadas por eletroforese, utilizando-se gel de agarose. Formaram-se grupos em relação à idade, ao sexo e à condição reprodutiva. Os valores médios das frações séricas albumina (2,85±0,36g/dL), alfa 1 (0,28±0,11g/dL), alfa 2 (0,26±0,08g/dL), beta 1 (0,57±0,15g/dL), beta 2 (0,89±0,28g/dL), gamaglobulinas (1,86±0,34g/dL), relação albumina/globulina (0,75±0,18) e os percentis 2,5% e 97,5% apresentaram diferenças pontuais em relação aos intervalos propostos para a espécie. Observaram-se maiores valores de alfa 1, alfa 2 globulinas, no grupo de seis a oito anos, e de gamaglobulinas, no grupo acima de 13 anos de idade. O proteinograma sérico foi similar entre machos e fêmeas e entre fêmeas vazias e gestantes. Sexo e estado gestacional não afetaram o proteinograma sérico. Alfa e gamaglobulinas têm incrementos em função de idades crescentes. O proteinograma sérico de equinos Campeiros tem variações que devem ser consideradas em exames laboratoriais.(AU)

Serum amyloid A and plasma protein electrophoresis fractions in farmed white-tailed deer.

Tools to measure the acute-phase response have been utilized widely in veterinary medicine. Evaluation by plasma protein electrophoresis (PPEP) has become an increasingly common assay in veterinary clinical pathology. Commercial reagents for serum amyloid A (SAA) have been validated for use in a variety of wildlife species. We analyzed samples from 29 healthy fawns and 60 healthy adult farmed white-tailed deer (WTD; Odocoileus virginianus) using an automated assay for SAA and a semi-automated method for PPEP. The robust statistical method for reference interval generation was used. SAA levels in fawns (0.1-26 mg/L) were found to be significantly higher than those in adults (0.1-5 mg/L, p < 0.01). The mean total protein was significantly lower in fawns (48 ± 10 g/L, p < 0.01) than in adults (73 ±5 g/L). The albumin-to-globulin ratio was also lower in fawns (0.56 ± 0.14) than in adults (1.25 ± 0.19, p < 0.01). Changes in SAA levels were observed in a variety of clinically abnormal animals. The combined use of the automated and semi-automated assays in our study may provide an additional valuable assessment tool in the care of captive WTD populations, for research studies, and for monitoring free-ranging animals.

Preliminary data on possible protein markers of parturition in cows.

Parturition is one of the most important events in reproduction. Regardless of many studies, exact time for pregnancy termination and onset of parturition is impossible to determine. The aim of this study was to describe and to compare protein profile of plasma from healthy pregnant cows (n = 6) at following five time points: 2 weeks, 1 week before, at parturition, 1 week and 2 weeks after parturition to search for possible protein markers of parturition. Plasma samples were analysed by 1D and 2D electrophoresis, and selected spots were identified by mass spectrometry. Protein profile showed no uniform pattern. Seventy spots differed at least for one sampling point from the time point 2 weeks before parturition which served as reference. Thirty spots expressed higher intensity of staining 1 week as 2 weeks before parturition while 13 showed opposite relationship. Twenty two spots expressed higher intensity of staining at parturition as 2 weeks before delivery while 15 showed opposite relationship. Eighteen spots expressed higher intensity of staining 2 weeks before parturition as 1 week post-partum while 2 showed opposite relationship. Fifteen spots expressed higher intensity of staining 2 weeks before parturition as 2 weeks after delivery while 14 showed opposite relationship. Thirty-five proteins, belonging to different functional groups, were identified. Of them, 15 spots differed significantly between parturition and 2 weeks before delivery. Among them were metalloproteinase inhibitor and LDH which seem to be the most promising molecules considered as parturition markers due to their functions.

Serum levels of proteins and acute-phase proteins in captive emus (Dromaius novaehllandieae) of different ages / Teores séricos de proteínas, inclusive proteínas de fase aguda, em emus (Dromaius novaehollandiae) de diferentes faixas etárias e criados em cativeiro

Protein electrophoresis is a relatively simple technique that allows separating serum protein fractions, and provides important information in the investigation and diagnosis of several diseases. This study determined the levels of acute-phase proteins in the serum of healthy, captive emus (Dromaius novaehollandiae). Animals were divided into two groups (n=11 in each) based on age, with 1-year-old and 4-year-old emus. Acute-phase proteins were separated by SDS-PAGE. Ceruloplasmin, transferrin, albumin, haptoglobin, acidic glycoprotein, IgA, and IgG were detected in the serum of all animals. Protein profiles varied significantly with age (P<0.05). Individuals in the 4-year-old emus group had higher values of ceruloplasmin, transferrin, albumin, haptoglobin, and acidic glycoprotein, compared with the group with 1-year-old animals, showing the role of age in the protein profile of this species. Reference values for acute-phase proteins in healthy emus may be useful in the evaluation of health status and in the diagnosis of diseases affecting the species.(AU)

A eletroforese de proteínas é um método relativamente simples, que permite a separação das proteínas do plasma em frações. Sua interpretação fornece informações importantes para a investigação e o diagnóstico de inúmeras doenças. O objetivo deste estudo foi o de determinar a concentração das proteínas de fase aguda no soro de emus (Dromaius novaehollandiae) hígidos e criados em cativeiro. As aves foram separadas em dois grupos: grupo 1: (n=11), aves com um ano de idade; grupo 2: (n=11), aves com quatro anos de idade. As proteínas de fase aguda foram separadas por eletroforese em gel de poliacrilamida (SDS-PAGE). Identificaram-se as proteínas ceruloplasmina, transferrina, albumina, IgG, haptoglobina, glicoproteína ácida, IgA e IgG no soro de todos os emus. Houve diferença (P<0.05) entre os traçados eletroforéticos em função da faixa etária. As aves do grupo 2 apresentaram valores superiores de ceruloplasmina, transferrina, albumina, haptoglobina e glicoproteína ácida quando comparadas às aves do grupo 1. Conclui-se que o perfil eletroforético de emus sofre alterações conforme a idade analisada. O estabelecimento de valores de referência para as proteínas de fase aguda de emus hígidos poderá auxiliar estudos futuros na avaliação da saúde assim como no diagnóstico de doenças em emus.(AU)

EFFECTS OF "SWIM WITH THE TURTLES" TOURIST ATTRACTIONS ON GREEN SEA TURTLE (CHELONIA MYDAS) HEALTH IN BARBADOS, WEST INDIES.

Along the West Coast of Barbados a unique relationship has developed between endangered green sea turtles (Chelonia mydas) and humans. Fishermen began inadvertently provisioning these foraging turtles with fish offal discarded from their boats. Although initially an indirect supplementation, this activity became a popular attraction for visitors. Subsequently, demand for this activity increased, and direct supplementation or provisioning with food began. Food items offered included raw whole fish (typically a mixture of false herring [Harengula clupeola] and pilchard [Harengula humeralis]), filleted fish, and lesser amounts of processed food such as hot dogs, chicken, bread, or various other leftovers. Alterations in behavior and growth rates as a result of the provisioning have been documented in this population. The purpose of this study was to determine how tourism-based human interactions are affecting the overall health of this foraging population and to determine what potential health risks these interactions may create for sea turtles. Juvenile green sea turtles (n=29) were captured from four sites off the coast of Barbados, West Indies, and categorized into a group that received supplemental feeding as part of a tour (n=11) or an unsupplemented group (n=18) that consisted of individuals that were captured at sites that did not provide supplemental feeding. Following capture, a general health assessment of each animal was conducted. This included weight and morphometric measurements, a systematic physical examination, determination of body condition score and body condition index, epibiota assessment and quantification, and clinical pathology including hematologic and biochemical testing and nutritional assessments. The supplemented group was found to have changes to body condition, vitamin, mineral, hematologic, and biochemical values. Based on these results, recommendations were made to decrease negative behaviors and health impacts for turtles as a result of this provisioning.

Protein and cholesterol electrophoresis of plasma samples from captive cownose ray (Rhinoptera bonasus).

Our study was undertaken to assess the application of semiautomated methods available at the reference laboratory level for the evaluation of plasma protein and cholesterol via electrophoresis in samples from cownose rays (Rhinoptera bonasus). Three groups of animals were assessed: clinically normal, clinically abnormal, and parasitized with leeches. As reported previously, the albumin band was negligible; the protein electrophoretograms were dominated by a large beta-globulin fraction. While the group of samples from the leech-parasitized rays did not show any large differences, the abnormal group exhibited significantly elevated total solids and cholesterol levels. The latter was related to a significant increase in very low density lipoprotein levels. The results demonstrate the potential application of these laboratory methods in quantitation of plasma proteins and cholesterol fractions in subclass Elasmobranchii.