Hepatocyte paraffin 1 (Hep Par 1), a murine monoclonal antibody, is widely used in surgical pathology practice to determine the hepatocellular origin of neoplasms. However, identity of the antigen for Hep Par 1 is unknown. The aim of this study was to characterize the Hep Par 1 antigen. To identify the antigen, immunoprecipitation was used to isolate the protein from human liver tissue, and a distinct protein band was detected at approximately 165 kDa. The protein band was also present in small intestinal tissue, but was not present in several other non-liver tissues nor in three human hepatocellular carcinoma cell lines, Huh-7, HepG2, and LH86. The protein was purified and analyzed by mass spectrometry. It was identified as carbamoyl phosphate synthetase 1 (CPS1). CPS1 is a rate-limiting enzyme in urea cycle and is located in mitochondria. We demonstrated that hepatoid tumors (gastric and yolk sac) were immunoreactive with both Hep Par 1 antibody and anti-CPS1 antibody, further confirming the results of mass spectrometric analysis. We found that the three human hepatocellular carcinoma cell lines do not express either CPS1 RNA or protein. We confirmed that the gene was present in these cell lines, suggesting that suppression of CPS1 expression occurs at the transcriptional level. This finding may have relevance to liver carcinogenesis, since poorly differentiated hepatocellular carcinomas exhibit poor to absent immunoreactivity to Hep Par 1. In conclusion, we have identified the antigen for Hep Par 1 antibody as a urea cycle enzyme CPS1. Our results should encourage further investigation of potential role that CPS1 expression plays in liver pathobiology and carcinogenesis. The histological distinction between hepatocellular carcinomas (HCC) and metastatic adenocarcinoma to the liver can sometimes be a challenging dilemma for surgical pathologists, particularly given the histological variants of HCC that can occur. In addition, tumors in other sites can display hepatoid morphologic features, adding to the diagnostic challenge when considering their metastasis to the liver. In the end, a wide panel of immunohistochemical markers is often used for the differential diagnosis of HCC, cholangiocarcinoma and metastatic adenocarcinoma. These markers include alpha-fetoprotein (AFP), polyclonal carcinoembryonic antigen (pCEA), and alpha-1-antitrypsin.

Protonated molecular peptide ions and their product ions generated by tandem mass spectrometry appear as isotopologue clusters due to the natural isotopic variations of carbon, hydrogen, nitrogen, oxygen and sulfur. Quantitation of the isotopic composition of peptides can be employed in experiments involving isotope effects, isotope exchange, isotopic labeling by chemical reactions, and studies of metabolism by stable isotope incorporation. Both ion trap and quadrupole-time of flight mass spectrometry are shown to be capable of determining the isotopic composition of peptide product ions obtained by tandem mass spectrometry with both precision and accuracy. Tandem mass spectra obtained in profile-mode of clusters of isotopologue ions are fit by non-linear least squares to a series of Gaussian peaks (described in the accompanying manuscript) which quantify the M n / M 0 values which define the isotopologue distribution (ID). To determine the isotopic composition of product ions from their ID, a new algorithm that predicts the M n /M 0 ratios is developed which obviates the need to determine the intensity of all of the ions of an ID. Consequently a precise and accurate determination of the isotopic composition a product ion may be obtained from only the initial values of the ID, however the entire isotopologue cluster must be isolated prior to fragmentation. Following optimization of the molecular ion isolation width, fragmentation energy and detector sensitivity, the presence of isotopic excess ( 2 H, 13 C, 15 N, 18 O) is readily determined within 1%. The ability to determine the isotopic composition of sequential product ions permits the isotopic composition of individual amino acid residues in the precursor ion to be determined. Keywordsisotopologue distribution; mass isotopomer distribution; tandem mass spectrometry; deuterium incorporation; isotopic excess; isotope quantitation; H/D exchange; protein turnover Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. Stable isotopic labels can be incorporated randomly into proteins and peptides through chemical exchange, metabolic labeling with tracers, and through chemical modifications used for quantitation in proteomic studies. The quantitation of isotopically modified proteins and peptides is necessary for all of these applications. Tandem mass spectrometry results in higher specificity and sensitivity for many analytical purposes. As routinely applied, information on the isotopic composition of the peptides is routinely lost due to the isolation of only the base ion during tandem mass spectrometry. We demonstrate th...

Haloacetic acids (HAAs) are environmentally and medically important chemicals. No analytical method is currently available to analyze EPA-regulated HAAs in biological samples at environmentally relevant low concentrations. Clinical studies of this class of chemicals are also limited by the lack of analytical techniques of high sensitivity and precision. We now report a new analytical method using gas chromatography/ion trap mass spectrometry for quantifying nine HAAs present inplasma, urine, and water at picogram per milliliter levels. The derivatization reactions of HAAs with pentafluorobenzyl bromide were optimized and detection with an electron capture negative ion chemical ionization mode was employed to enhance the sensitivity. Selected ion monitoring and selected reaction monitoring methods were utilized for quantitation. The detection limits of HAAs in plasma, urine, and water were 25-1000 pg/mL. Accuracies varied from 86.6 to 118.1% (intraday) and 81.7 to 119.6% (interday). Precisions (CV) varied from 0.9 to 19.9% (intraday) and 0.8 to 19.8% (interday), and linearities (r2) varied from 0.9732 to 0.9998 (intraday) and 0.9422 to 0.9987 (interday), respectively. Methyl tertbutyl ether and diethyl ether provided the highest extraction recoveries for the HAAs (74.9-107.2%). The method was applied successfully to a kinetic investigation of low levels of HAAs in humans consuming chlorinated drinking water.

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Mesoporous silica nanoparticles (MSN) are widely used as pesticide carriers to enhance their effective utilization, since it can promote the solubility and absorption of pesticides by plants. For plants, the particle size of pesticides influences their absorption and efficacy. Herein, is our research work of the size effect of MSN on the loading, release, and delivery behavior of pyraoxystrobin (Pyr) in cucumber plants. The well-ordered Pyr-loaded carbon quantum dots-MSN (Pyr@M) with sizes of 15, 100, and 200 nm were prepared. A comparative study among different particle sizes of Pyr@M was carried out on the aspects of control release performance, loading content, uptake, and transportation performance in cucumber plants. It was found that the loading content increased as the particle size increased. The nanoparticles as carriers increased the solubility of insoluble Pyr, but the nanoparticle size had no clear difference impact on the release rate. The efficiency of the cellular uptake strongly depended on the particle size. The smaller the MSN size, the easier it was to be absorbed and transmitted by cucumber plants. Compared to the free Pyr, the upward transportation rate of Pyr from Pyr@M in plant increased by 3.5 times. These findings provide new theoretical basis to design the MSN pesticide delivery system.

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