scholarly journals Optimized liquid and gas phase fractionation increase HLA-peptidome coverage for primary cell and tissue samples

2021 ◽  
Author(s):  
Susan Klaeger ◽  
Annie Apffel ◽  
Karl R Clauser ◽  
Siranush Sarkizova ◽  
Giacomo Oliveira ◽  
...  
Keyword(s):  
Ion Mobility ◽  
Reversed Phase ◽  
Current Standard ◽  
Binding Prediction ◽  
High Coverage ◽  
Tissue Samples ◽  
Input Material ◽  
Increased Sensitivity ◽  
Wet Weight ◽  
Hla Binding

Mass spectrometry is the most effective method to directly identify peptides presented on HLA molecules. However, current standard approaches often require many millions of cells for input material to achieve high coverage of the immunopeptidome and are therefore not compatible with the often limited amounts of tissue available from clinical tumor samples. Here, we evaluated microscaled basic reversed-phase fractionation to separate HLA peptide samples off-line followed by ion mobility coupled to LC-MS/MS for analysis. The combination of these two separation methods enabled identification of 20% to 50% more peptides compared to samples analyzed without either prior fractionation or use of ion mobility alone. We demonstrate coverage of HLA immunopeptidomes with up to 8,107 distinct peptides starting with as few as 100 million cells or 150 milligrams of wet weight tumor tissue. This increased sensitivity can improve HLA binding prediction algorithms and enable detection of clinically relevant epitopes such as neoantigens.

scholarly journals Prophylactic impact of nano-selenium on performance, carcasses quality, and tissues' selenium concentration using reversed-phase high-performance liquid chromatography during microbial challenge in broiler chickens

2020 ◽  
Vol 13 (9) ◽  
pp. 1780-1797
Author(s):  
Essam S. Soliman ◽  
Fadwa F. Mahmoud ◽  
Mai A. Fadel ◽  
Rania T. Hamad
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
Meat Quality ◽  
High Performance ◽  
Bacterial Load ◽  
Reversed Phase ◽  
Bursa Of Fabricius ◽  
Health Concern ◽  
Tissue Architecture ◽  
Tissue Samples

Background and Aim: Nano-selenium (NS) supplementation contributes in improving productivity, performance, and meat quality while reducing public health concern. Influence of NS and inorganic selenium (Se) water additive on performance, carcass quality, immunoglobulin concentration, intestinal microbiota, Se tissue concentrations, and tissue architecture was studied. Materials and Methods: Two-hundred and sixty 1-day-old Hubbard chicks were randomly grouped into five groups (5×52) and supplemented with 0.5 and 1.0 mL of NS and inorganic Se (100 mg.L-1). G1, G2, G3, and G4 were challenged with Escherichia coli O157: H7 2.6×108 on the 14th day. A total of 2250 samples, including 250 sera, 250 intestinal swabs, and 1500 organ and tissue samples as liver, spleen, heart, bursa, intestine, and breast muscles, and 250 eviscerated carcasses were collected. Results: The results revealed a highly significant increase (p<0.01) in live body weights, weight gains, performance indices, carcasses, and organs weights, whereas immunoglobulin G and M concentrations in broilers treated with 0.5 and 1.0 mL NS, respectively, synchronized reveal a highly significant decline (p<0.01) in total bacterial and Enterobacteriaceae counts of intestinal swabs and breast muscles, final pH24, and drip loss in broilers treated with 0.5 and 1.0 mL NS, respectively. Meanwhile, water holding capacity revealed no significant differences between all groups. Reversed-phase high-performance liquid chromatography examination revealed the earlier disappearance of NS residues than inorganic Se from the broiler's liver and muscles. Histopathological photomicrographs of the liver, spleen, bursa of Fabricius, and intestine, as well as, the immunohistochemistry of intestinal sections revealed superior tissue architecture in broilers treated with NS contrary to inorganic Se. Conclusion: The study showed significant stimulation actions of NS on performance, immunity, carcass and meat quality, intestinal and muscles' bacterial load as well as short withdrawal period and nearly normal cellular architecture compared to inorganic Se.

Catecholamine measurements by high-performance liquid chromatography

1984 ◽  
Vol 247 (1) ◽  
pp. E13-E20 ◽  
Author(s):  
P. Hjemdahl
Keyword(s):  
High Performance Liquid Chromatography ◽  
Liquid Chromatography ◽  
Cation Exchange ◽  
High Performance ◽  
Reversed Phase ◽  
Exchange Chromatography ◽  
Lower Sensitivity ◽  
Tissue Samples ◽  
Basal Plasma ◽  
Postcolumn Derivatization

The development of sensitive detectors has allowed the use of high-performance liquid chromatography (HPLC) for measurements of catecholamines in extracts of plasma, urine, and tissue samples. Separation of the catecholamines may be effected by reversed phase chromatography or cation-exchange chromatography and quantitation by electrochemical detection (EC) or by fluorometry coupled with postcolumn derivatization according to the trihydroxyindole (THI) method. EC has a somewhat lower sensitivity than the THI method for norepinephrine (NE) and epinephrine (E). The THI method is insensitive to dopamine (DA). Basal plasma E levels of 0.1 nM (20 pg/ml) or less may be measured in sample volumes of 1-2 ml with EC. Sensitivity and reproducibility of an assay is not necessarily a guarantee of accuracy. It is argued that new methods and modifications of old methods should be validated against accepted methodology. This is rarely the case. Cation exchange HPLC with EC has been adequately validated, but only one of the reversed phase methods has been compared with radioenzymatic methodology. HPLC has the advantages of economy, speed, and more stimulating laboratory work, as compared with radioenzymatic methodology.

scholarly journals Ion mobility spectrometer with orthogonal X-Ray source for increased sensitivity

Talanta ◽  
2018 ◽  
Vol 185 ◽  
pp. 537-541 ◽  
Author(s):  
Erik Bunert ◽  
Tobias Reinecke ◽  
Ansgar T. Kirk ◽  
Alexander Bohnhorst ◽  
Stefan Zimmermann
Keyword(s):  
Ion Mobility ◽  
X Ray ◽  
Ion Mobility Spectrometer ◽  
Increased Sensitivity

scholarly journals Polycyclic aromatic hydrocarbons in porcine and bovine organs and tissues

2012 ◽  
Vol 51 (No. 5) ◽  
pp. 239-247 ◽  
Author(s):  
M. Ciganek ◽  
J. Neca
Keyword(s):  
Adipose Tissue ◽  
Polycyclic Aromatic Hydrocarbons ◽  
Aromatic Hydrocarbons ◽  
Vitreous Humour ◽  
Tissue Samples ◽  
Polycyclic Aromatic ◽  
Wet Weight ◽  
Pah Exposure ◽  
Animal Exposure ◽  
Kidney Liver

Concentrations of 16 polycyclic aromatic hydrocarbons (PAH) were determined in porcine and bovine kidney, liver, lung, muscle and adipose tissue samples, and in eyeballs (lens and vitreous humour) in fattener pigs and cows. The total average PAH concentrations in individual organs were: 5.4, 6.3 (kidney); 3.8, 2.7 (liver); 4.6, 5.4 (lung); 3.6, 5.1 (muscle tissue); 0.05, 0.11 (adipose tissue); 57.9, 16.3 (lens) and 14, 6.4 (vitreous humour) for pigs and cows in ng/g of wet weight, respectively. Phenanthrene, naphthalene, pyrene and fluoranthene were predominant PAH present in samples. No significant differences (P &gt; 0.05) were found among distribution of PAH in animal bodies from several localities with various PAH exposure or between their levels in porcine and bovine organs and tissues, except for eyeballs. On the contrary, significant variations of PAH concentrations (P&nbsp;&lt;&nbsp;0.01) were found between species in the same tissues from the same stable. The highest total concentrations of PAH were found in porcine and bovine lenses. Analyses of porcine and/or bovine lenses for PAH content could be used for determination of animal exposure to these compounds.

The detection of AKH/HrTH-like peptides inAscaridia galliandAscaris suumusing an insect hyperglycaemic bioassay

Parasitology ◽  
1994 ◽  
Vol 108 (4) ◽  
pp. 479-485 ◽  
Author(s):  
T. R. B. Davenport ◽  
L. A. Eaves ◽  
T. K. Hayes ◽  
D. L. Lee ◽  
R. E. Isaac
Keyword(s):  
High Performance ◽  
Periplaneta Americana ◽  
Reversed Phase ◽  
Parasitic Nematodes ◽  
Ascaridia Galli ◽  
Adipokinetic Hormone ◽  
Blaberus Discoidalis ◽  
Peptide Fraction ◽  
Wet Weight ◽  
Dose Dependent

SUMMARYEvidence for the presence of adipokinetic hormone/hypertrehalosaemic hormone (AKH/HrTH)-like peptides in the parasitic nematodesAscaridia galliandAscaris suumhas been obtained using insect bioassays which measure hyperglycaemic responses to peptides belonging to the AKH/HrTH family of insect hormones. A peptide fraction extracted from heads and tails ofAscaridia gallievoked a dose-dependent hyperglycaemic response when injected into the cockroach,Periplaneta americana. Maximal bioactivity was obtained with material that was equivalent to 38 mg (wet weight) of nematode. Bioactivity appeared to be highest in extracts from heads and tails of both male and female worms and could be fractionated into at least three peaks of hyperglycaemic activity by reversed-phase high-performance liquid chromatography. An extract from heads and tails ofA. suumalso evoked a hyperglycaemic response when injected into the cockroach,Blaberus discoidalis. The bioactivity was inactivated on incubation with pure endopeptidase 24.11, confirming the peptidic nature of the bioactive material. These results provide evidence for the existence of peptides related to the insect AKH/HrTH family of peptides in parasitic nematodes.

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