Method Validation and Establishment of Reference Intervals for an Insulin-like Growth Factor-1 Chemiluminescent Immunoassay in Cats

Previously, radioimmunoassay (RIA) has been the only assay to measure insulin-like growth factor-1 (IGF-1) to diagnose hypersomatotropism (HS). Due to radiation concerns, availability, and the cost of IGF-1 RIA, validation of assays for automated analysers such as a chemiluminescent immunoassay (CLIA) is needed. The aim of this study was to validate a CLIA for measurement of feline IGF-1 (IMMULITE 2000® XPi, Siemens Medical Solutions Diagnostics, Malvern, PA, USA) compared to IGF1 RIA, establish reference interval (RI), and determine a cut-off value for diagnosis of HS in diabetic cats. Validation of assay performance included precision, linearity, and recovery studies. Right-sided RI was determined using surplus serum of 50 healthy adult cats. Surplus serum samples of diabetic cats with known IGF-1 concentration with (n = 32/68) and without HS (n = 36/68) were used for method comparison with RIA. The cut-off for diagnosis of HS was established using receiver operating characteristic (ROC) analysis. The intra-assay coefficient of variation (CV) was ≤4.7%, and the inter-assay CV was ≤5.6% for samples with low, medium, and high IGF-1 concentration. Linearity was excellent (R2 > 0.99). The correlation between CLIA and RIA was very high (rs = 0.97), with a mean negative bias for CLIA of 24.5%. The upper limit of RI was 670 ng/mL. ROC analysis showed an area under the curve of 0.94, with best cut-off for diagnosis of HS at 746 ng/mL (sensitivity, 84.4%; specificity, 97.2%). The performance of CLIA was good, and the RI and cut-off for HS diagnosis established in this study allow for CLIA to be used in routine work-up of diabetic cats

Odor Determination at Wastewater Collection Systems: Olfactometry versus H2S Analyses

H2S is usually the main indicator parameter to assess when nuisance odors arise in wastewater handling systems. However, sampling and measurement of H2S need to be made with utmost care and analytical methods are neither easy nor cheap. Although Jerome meters from Arizona Instruments are used in some parts of the world, the cost and difficulty of measuring H2S in odorous gases is a general problem. It should also be kept in mind that a simple linear relationship between H2S concentrations and odor perception by the public is not available, especially in complex odorous gas mixtures. This leads to the conclusion that measuring the H2S alone is neither easy nor accurate in describing and resolving public complaints due to odors from wastewater collection and treatment systems. In this study, data for odors and H2S measurements from two pumping stations of the wastewater collection system of Cesme, Izmir, Turkey are evaluated. Concentrations of H2S were measured with chemical methods. Results indicated concentrations several orders of magnitude above the odor threshold level, which is generally accepted as 0.1 mu g/m(3) for pure H2S in air. The statistical relationships between analytical and sensory odor data were established. Correlations indicated power law relationships between odor and H2S concentrations (R-2 between 0.88-0.92). Steven's law between the intensity of human perception and concentration of stimulants is obeyed

Figures for Pituitary pathology and gene expression in acromegalic cats

<div> <p>Figure 1</p> <p>Representative photomicrographs of growth hormone (A - C) and prolactin (D – F) immunostaining. A and D are x40 photomicrographs demonstrating specific immunostaining for somatrophs and lactotrophs, respectively. B and E are photomicrographs from a control cat C and F are from an acromegalic cat. </p><p><br></p> <p> </p> <p>Figure 2</p> <p>Representative images of SSTR2 immunoreactivity using feline pituitary tissue. A - D represent pituitary tissue exhibiting SSTR2 immunohistochemistry scores 0, 1, 2 and 3, respectively using the following criteria:.0 = absent; 1 = cytoplasmic staining; 2 = membranous staining in less than 50% cells or incomplete membranous staining; and 3 = circumferential membranous staining in >50% cells. All presented photomicrographs collected at x100 magnification using Leica DM400 B, Leica Microsystems Cambridge, UK.</p><p><br></p> <p> </p> <p>Figure 3</p> <p>An electropherogram results from PCR products using multiplex 1 primer sets The blue peaks represent PCR products from gene specific primers and red peaks represent product size standards.</p><p><br></p> <p> </p> <p>Figure 4</p> <p>All images stained using Silver stain for reticulin fibres and counter stained using Nuclear Fast Red solution. A and C; reconstructed stitched pituitary x100 magnification photomicrographs from two control pituitaries. B and D; x400 magnification photomicrographs from A and C, respectively. The acinar pattern of reticulin staining is identified in B and D. This pattern of reticulin staining was demonstrated in all reticulin staining control pituitaries.</p><p><br></p> <p> </p> <p>Figure 5</p> <p>All images stained using Silver stain for reticulin fibres and counter stained using Nuclear Fast Red solution. A – D; selected images taken from reconstructed stitched pituitary x100 magnification photomicrographs from four HST pituitaries. A; disrupted reticulin staining and loss of acinar structure. B; areas of enlarged acini (blue stars) and areas of loss of acinar structure (blue cross). C; enlarged acini (blue stars) adjacent to normal sized and small acini. D; loss of acinar structure in the bottom right of the image (blue stars), and adenomatous tissue has compressed the normal pituitary tissue resulting in compression of the acini and a ring of cords of acini giving the impression of a pseudocapsule.</p><p><br></p> <p> </p> <p>Figure 6</p> <p>A: Bar charts comparing the relative gene expression of <i>SSTR1</i>, <i>SSTR2 and</i> <i>SSTR5</i> in pituitary tissue from control (CTRL) and acromegalic (Acro) cats determine using GeXP multiplex technique. <i>RPL18</i> is the reference gene. Bar height represents mean and error bars are 95% confidence intervals ** represents <i>P</i> < 0.01 and *** represents <i>P</i> < 0.001. Dot plot of the individual somatostatin profiles from each of the 19 acromegalic cats.</p> </div> <br> <p>Table 1</p> <p>Clinical data of cats in the control and acromegalic groups</p><p><br></p> <p> </p> <p>Table 2</p> <p>Gene expression data and GH, PRL and SSTR2 immunohistochemistry scoring of cats in the control and acromegalic groups.</p><p><br></p> <p> </p> <p>Table 3</p> <p>Summary of Spearman rank correlation gene expression data in the control group and acromegalic groups</p><p><br></p> <p> </p><p>Supplementary material 1</p><p>Nucleotide sequences of primers used in multiplexes 1, 2 and 3.</p><p><br></p><p>Supplementary material 2</p><p>Amino acid multiple sequence alignment for human and feline SSTR2</p><div><br></div><p>Supplementary material 3</p><p>Amino acid multiple sequence alignment for porcine and feline growth hormone </p><div><br></div><p>Supplementary material 4</p><p>Amino acid multiple sequence alignment for porcine and feline prolactin </p><div><br></div><div><br></div