Toward Label-Free Selective Cell Separation of Different Eukaryotic Cell Lines Using Thermoresponsive Homopolymer Layers

2019 ◽  
Vol 2 (6) ◽  
pp. 2557-2566 ◽  
Author(s):  
Siyu Jiang ◽  
Mareike Müller ◽  
Holger Schönherr
Keyword(s):  
Cell Lines ◽  
Cell Separation ◽  
Eukaryotic Cell ◽  
Label Free

Assay for Adherence of Vibrio cholerae to Eukaryotic Cell Lines

BIO-PROTOCOL ◽  
2014 ◽  
Vol 4 (8) ◽  
Author(s):  
Amit Dey ◽  
Abha Bhagat ◽  
Rukhsana Chowdhury
Keyword(s):  
Vibrio Cholerae ◽  
Cell Lines ◽  
Eukaryotic Cell

scholarly journals In Vitro Spectroscopy-Based Profiling of Urothelial Carcinoma: A Fourier Transform Infrared and Raman Imaging Study

Cancers ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 123
Author(s):  
Monika Kujdowicz ◽  
Wojciech Placha ◽  
Brygida Mech ◽  
Karolina Chrabaszcz ◽  
Krzysztof Okoń ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
Fourier Transform ◽  
Cancer Cells ◽  
Cell Line ◽  
Cell Lines ◽  
Malignant Neoplasm ◽  
Fourier Transform Infrared ◽  
Diagnostic Tools ◽  
Label Free ◽  
Bladder Cancer Cells

Markers of bladder cancer cells remain elusive, which is a major cause of the low recognition of this malignant neoplasm and its recurrence. This implies an urgent need for additional diagnostic tools which are based on the identification of the chemism of bladder cancer. In this study, we employed label-free techniques of molecular imaging—Fourier Transform Infrared and Raman spectroscopic imaging—to investigate bladder cancer cell lines of various invasiveness (T24a, T24p, HT-1376, and J82). The urothelial HCV-29 cell line was the healthy control. Specific biomolecules discriminated spatial distribution of the nucleus and cytoplasm and indicated the presence of lipid bodies and graininess in some cell lines. The most prominent discriminators are the total content of lipids and sugar moieties as well as the presence of glycogen and other carbohydrates, un/saturated lipids, cytochromes, and a level of S-S bridges in proteins. The combination of the obtained hyperspectral database and chemometric methods showed a clear differentiation of each cell line at the level of the nuclei and cytoplasm and pointed out spectral signals which differentiated bladder cancer cells. Registered spectral markers correlated with biochemical composition changes can be associated with pathogenesis and potentially used for the diagnosis of bladder cancer and response to experimental therapies.

Eukaryotic cell lines as a sensitive layer for direct monitoring of carbon monoxide

2011 ◽  
Vol 208 (6) ◽  
pp. 1345-1350 ◽  
Author(s):  
Ulrich Bohrn ◽  
Evamaria Stütz ◽  
Maximilian Fleischer ◽  
Michael J. Schöning ◽  
Patrick Wagner
Keyword(s):  
Carbon Monoxide ◽  
Cell Lines ◽  
Eukaryotic Cell ◽  
Sensitive Layer

Immunofluorescent localization of the ubiquitin-activating enzyme, E1, to the nucleus and cytoskeleton

1993 ◽  
Vol 264 (1) ◽  
pp. C93-C102 ◽  
Author(s):  
J. S. Trausch ◽  
S. J. Grenfell ◽  
P. M. Handley-Gearhart ◽  
A. Ciechanover ◽  
A. L. Schwartz
Keyword(s):  
Cell Lines ◽  
Chinese Hamster ◽  
Eukaryotic Cell ◽  
Nuclear Staining ◽  
Immunofluorescent Localization ◽  
Acid Protein ◽  
Ubiquitin Conjugation ◽  
Double Label ◽  
Pleiotropic Functions ◽  
Amino Acid Protein

Ubiquitin, a 76-amino acid protein, is covalently attached to abnormal and short-lived proteins, thus marking them for ATP-dependent proteolysis in eukaryotic cells. Ubiquitin is found within the cytoplasm, nucleus, microvilli, autophagic vacuoles, and lysosomes. The ubiquitin-activating enzyme, E1, catalyzes the first step in ubiquitin conjugation. To date, very little is known about the subcellular distribution of this enzyme. We have utilized immunofluorescence and immunoblotting to examine the cellular distribution of E1 in several eukaryotic cell lines, including HeLa, smooth muscle A7r5, choriocarcinoma BeWo, Pt K1, and Chinese hamster ovary (CHO) E36. E1 was identified in both cytoplasmic and nuclear compartments in all cell lines examined. However, the relative abundance within these compartments differed markedly between the cell lines. Even within a single cell line, nuclear distribution was not uniform, and certain cells demonstrated an absence of nuclear staining. E1 resides predominantly within the nucleus in BeWo. In contrast, its distribution in CHO and Pt K1 cells is mainly cytoplasmic. Within the cytoplasm, three pools of E1 were identified by double-label immunofluorescence. The first of these colocalized with phalloidin, indicating association of E1 with actin filaments. A second cytoplasmic pool colocalized with tubulin and was predominantly perinuclear in its distribution. The third pool associated with intermediate filaments. This suggests that E1 is associated with all three components of the cytoskeleton. The distribution of E1 was unaltered in a mutant line of CHO E36 designated ts20, in which the E1 can be thermally inactivated. The variable distribution of E1 among cell lines, including its apparent cytoskeletal association, suggests pleiotropic functions of this enzyme and the ubiquitin-conjugating system.

scholarly journals Secretomic Analysis Identifies Alpha-1 Antitrypsin (A1AT) as a Required Protein in Cancer Cell Migration, Invasion, and Pericellular Fibronectin Assembly for Facilitating Lung Colonization of Lung Adenocarcinoma Cells

2012 ◽  
Vol 11 (11) ◽  
pp. 1320-1339 ◽  
Author(s):  
Ying-Hua Chang ◽  
Shu-Hui Lee ◽  
I-Chuang Liao ◽  
Shin-Huei Huang ◽  
Hung-Chi Cheng ◽  
...  
Keyword(s):  
Lung Adenocarcinoma ◽  
Cancer Cells ◽  
Cell Lines ◽  
Metastatic Potential ◽  
Cell Surface Receptor ◽  
Migration And Invasion ◽  
Secretory Proteins ◽  
Label Free ◽  
Altered Expression ◽  
Lung Colonization

Metastasis is a major obstacle that must be overcome for the successful treatment of lung cancer. Proteins secreted by cancer cells may facilitate the progression of metastasis, particularly within the phases of migration and invasion. To discover metastasis-promoting secretory proteins within cancer cells, we used the label-free quantitative proteomics approach and compared the secretomes from the lung adenocarcinoma cell lines CL1-0 and CL1-5, which exhibit low and high metastatic properties, respectively. By employing quantitative analyses, we identified 660 proteins, 68 of which were considered to be expressed at different levels between the two cell lines. High levels of A1AT were secreted by CL1-5, and the roles of A1AT in the influence of lung adenocarcinoma metastasis were investigated. Molecular and pathological confirmation demonstrated that altered expression of A1AT correlates with the metastatic potential of lung adenocarcinoma. The migration and invasion properties of CL1-5 cells were significantly diminished by reducing the expression and secretion of their A1AT proteins. Conversely, the migration and invasion properties of CL1-0 cells were significantly increased through the overexpression and secretion of A1AT proteins. Furthermore, the assembly levels of the metastasis-promoting pericellular fibronectin (FN1), which facilitates colonization of lung capillary endothelia by adhering to the cell surface receptor dipeptidyl peptidase IV (DPP IV), were higher on the surfaces of suspended CL1-5 cells than on those of the CL1-0 cells. This discovery reflects previous findings in breast cancer. In line with this finding, FN1 assembly and the lung colonization of suspended CL1-5 cells were inhibited when endogenous A1AT protein was knocked down using siRNA. The major thrust of this study is to demonstrate the effects of coupling the label-free proteomics strategy with the secretomes of cancer cells that differentially exhibit invasive and metastatic properties. This provides a new opportunity for the effective identification of metastasis-associated proteins that are secreted by cancer cells and promote experimental metastasis.

scholarly journals Phosphotyrosine-based Phosphoproteomics for Target Identification and Drug Response Prediction in AML Cell Lines

2020 ◽  
Vol 19 (5) ◽  
pp. 884-899 ◽  
Author(s):  
Carolien van Alphen ◽  
Jacqueline Cloos ◽  
Robin Beekhof ◽  
David G. J. Cucchi ◽  
Sander R. Piersma ◽  
...  
Keyword(s):  
Cell Lines ◽  
Tyrosine Kinases ◽  
Kinase Inhibitor ◽  
Target Identification ◽  
Response Prediction ◽  
Clinical Samples ◽  
Label Free ◽  
Poor Treatment ◽  
Poor Treatment Outcome ◽  
The Individual

Acute myeloid leukemia (AML) is a clonal disorder arising from hematopoietic myeloid progenitors. Aberrantly activated tyrosine kinases (TK) are involved in leukemogenesis and are associated with poor treatment outcome. Kinase inhibitor (KI) treatment has shown promise in improving patient outcome in AML. However, inhibitor selection for patients is suboptimal.In a preclinical effort to address KI selection, we analyzed a panel of 16 AML cell lines using phosphotyrosine (pY) enrichment-based, label-free phosphoproteomics. The Integrative Inferred Kinase Activity (INKA) algorithm was used to identify hyperphosphorylated, active kinases as candidates for KI treatment, and efficacy of selected KIs was tested.Heterogeneous signaling was observed with between 241 and 2764 phosphopeptides detected per cell line. Of 4853 identified phosphopeptides with 4229 phosphosites, 4459 phosphopeptides (4430 pY) were linked to 3605 class I sites (3525 pY). INKA analysis in single cell lines successfully pinpointed driver kinases (PDGFRA, JAK2, KIT and FLT3) corresponding with activating mutations present in these cell lines. Furthermore, potential receptor tyrosine kinase (RTK) drivers, undetected by standard molecular analyses, were identified in four cell lines (FGFR1 in KG-1 and KG-1a, PDGFRA in Kasumi-3, and FLT3 in MM6). These cell lines proved highly sensitive to specific KIs. Six AML cell lines without a clear RTK driver showed evidence of MAPK1/3 activation, indicative of the presence of activating upstream RAS mutations. Importantly, FLT3 phosphorylation was demonstrated in two clinical AML samples with a FLT3 internal tandem duplication (ITD) mutation.Our data show the potential of pY-phosphoproteomics and INKA analysis to provide insight in AML TK signaling and identify hyperactive kinases as potential targets for treatment in AML cell lines. These results warrant future investigation of clinical samples to further our understanding of TK phosphorylation in relation to clinical response in the individual patient.

scholarly journals Label-free in vitro toxicity and uptake assessment of citrate stabilised gold nanoparticles in three cell lines

2013 ◽  
Vol 10 (1) ◽  
pp. 50 ◽  
Author(s):  
Melissa A Vetten ◽  
Nonhlanhla Tlotleng ◽  
Delia Tanner Rascher ◽  
Amanda Skepu ◽  
Frankline K Keter ◽  
...  
Keyword(s):  
Gold Nanoparticles ◽  
Cell Lines ◽  
In Vitro Toxicity ◽  
Label Free

scholarly journals Double spiral microchannel for label-free tumor cell separation and enrichment

Lab on a Chip ◽  
2012 ◽  
Vol 12 (20) ◽  
pp. 3952 ◽  
Author(s):  
Jiashu Sun ◽  
Mengmeng Li ◽  
Chao Liu ◽  
Yi Zhang ◽  
Dingbin Liu ◽  
...  
Keyword(s):  
Tumor Cell ◽  
Cell Separation ◽  
Label Free ◽  
Double Spiral ◽  
Spiral Microchannel
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