scholarly journals Flow Cytometric Method for the Detection of Flavonoids in Cell Lines

2016 ◽  
Vol 21 (8) ◽  
pp. 858-865 ◽  
Author(s):  
Charlotte Grootaert ◽  
Gerard Bryan Gonzales ◽  
Hanne Vissenaekens ◽  
Tom Van de Wiele ◽  
Katleen Raes ◽  
...  
Keyword(s):  
Cell Lines ◽  
Cellular Uptake ◽  
Cellular Level ◽  
Animal Origin ◽  
Fixation Method ◽  
Future Research ◽  
Flavonoid Glycosides ◽  
Cell Type ◽  
Flow Cytometric Method ◽  
Flow Cytometric

Here, we describe an easy-to-use flow cytometric method using diphenylboric acid 2-amino ethyl ester (DPBA) stain for the detection of flavonoids in cells from human/animal origin. Flavonoid bioavailability and bioactivity depend on structure, conjugation and the cell type to which they are presented. We have studied cellular uptake of five flavonoids with different structures and conjugation forms. First, parameters including fixation method, technical and batch variability, and concentration were optimized. Second, uptake of two aglycones—quercetin and hesperetin—and their corresponding glycosides—rutin and hesperidin—in Caco-2 cells was compared. Third, the aglycone quercetin, glycoside rutin, and glucuronide baicalin were added to the Caco-2, HepG2, and CHO-K1 cell lines at 1, 10, and 20 µM concentrations and cellular uptake was measured after 1, 4, and 7 h. We conclude that quercetin was taken up by cells in a dose-dependent way, and that HepG2 cells had the highest uptake factors, followed by CHO-K1 and Caco-2 cells. Confocal microscopy showed cell type–dependent localization of quercetin in the cell membrane and cytoplasm. No uptake of flavonoid glycosides was detected. This flow cytometric method can be used for future research unravelling mechanisms behind flavonoid bioactivity in health and disease at the cellular level.

scholarly journals Establishment of HLA class I and MICA/B null HEK-293T panel expressing single MICA alleles to detect anti-MICA antibodies

2021 ◽  
Author(s):  
Ji-Ho Jeon ◽  
In-Cheol Baek ◽  
Cheol-Hwa Hong ◽  
Ki Hyun Park ◽  
Hyeyoung Lee ◽  
...  
Keyword(s):  
Cell Lines ◽  
Hla Class I ◽  
Cellular Level ◽  
Class I ◽  
Antibody Detection ◽  
Post Transplantation ◽  
Transplant Patients ◽  
Flow Cytometric Method ◽  
Complement Dependent Cytotoxicity

Abstract Pre- and post-transplantation anti-MICA antibody detection development are associated with an increased rejection risk and low graft survival. We previously generated HLA class I null HEK-293T using CRISPR/Cas9, while MICA and MICB genes were removed in this study. A panel of 11 cell lines expressing single MICA alleles was established. Anti-MICA antibody in the sera of kidney transplant patients was determined using FCM (flow cytometric method) and the Luminex method. In the 44 positive sera, the maximum FCM value was 2,879 MFI compared to 28,135 MFI of Luminex method. Eleven sera (25%) were determined as positive by FCM and 32 sera (72%) were positive by the Luminex method. The sum of total MICA antigens, MICA*002, *004, *009, *019, and *027 correlation showed a statistically significant between the two methods (P = 0.0412, P = 0.0476, P = 0.0019, P = 0.0098, P = 0.0467, and P = 0.0049). These results demonstrated that HEK-293T-based engineered cell lines expressing single MICA alleles were suitable for measuring specific antibodies against MICA antigens in the sera of transplant patients. Studies of antibodies to MICA antigens may help to understand responses in vivo and increase clinical relevance at the cellular level such as complement-dependent cytotoxicity.

A novel quantitative flow cytometric method for measuring glucocorticoid receptor (GR) in cell lines: Correlation with the biochemical determination of GR

2007 ◽  
Vol 324 (1-2) ◽  
pp. 110-119 ◽  
Author(s):  
Ioannis F. Voutsas ◽  
Angelos D. Gritzapis ◽  
Michael N. Alexis ◽  
Efrosini S. Katsanou ◽  
Sonia Perez ◽  
...  
Keyword(s):  
Glucocorticoid Receptor ◽  
Cell Lines ◽  
Flow Cytometric Method ◽  
Flow Cytometric ◽  
Quantitative Flow

Utilization of non-AUG initiation codons in a flow cytometric method for efficient selection of recombinant cell lines

2011 ◽  
Vol 108 (11) ◽  
pp. 2611-2622 ◽  
Author(s):  
Victor R. Cairns ◽  
Christine T. DeMaria ◽  
Francis Poulin ◽  
José Sancho ◽  
Ping Liu ◽  
...  
Keyword(s):  
Cell Lines ◽  
Flow Cytometric Method ◽  
Flow Cytometric ◽  
Recombinant Cell ◽  
Efficient Selection ◽  
Selection Of

scholarly journals Establishment of HLA class I and MICA/B null HEK-293T panel expressing single MICA alleles to detect anti-MICA antibodies

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ji-Ho Jeon ◽  
In-Cheol Baek ◽  
Cheol-Hwa Hong ◽  
Ki Hyun Park ◽  
Hyeyoung Lee ◽  
...  
Keyword(s):  
Cell Lines ◽  
Hla Class I ◽  
Cellular Level ◽  
Class I ◽  
Antibody Detection ◽  
Post Transplantation ◽  
Transplant Patients ◽  
Flow Cytometric Method ◽  
Complement Dependent Cytotoxicity

AbstractPre- and post-transplantation anti-MICA antibody detection development are associated with an increased rejection risk and low graft survival. We previously generated HLA class I null HEK-293T using CRISPR/Cas9, while MICA and MICB genes were removed in this study. A panel of 11 cell lines expressing single MICA alleles was established. Anti-MICA antibody in the sera of kidney transplant patients was determined using flow cytometric method (FCM) and the Luminex method. In the 44 positive sera, the maximum FCM value was 2879 MFI compared to 28,135 MFI of Luminex method. Eleven sera (25%) were determined as positive by FCM and 32 sera (72%) were positive by the Luminex method. The sum of total MICA antigens, MICA*002, *004, *009, *019, and *027 correlation showed a statistically significant between the two methods (P = 0.0412, P = 0.0476, P = 0.0019, P = 0.0098, P = 0.0467, and P = 0.0049). These results demonstrated that HEK-293T-based engineered cell lines expressing single MICA alleles were suitable for measuring specific antibodies against MICA antigens in the sera of transplant patients. Studies of antibodies to MICA antigens may help to understand responses in vivo and increase clinical relevance at the cellular level such as complement-dependent cytotoxicity.

Expression of Platelet Membrane Glycoprotein V in Human Megakaryocytes and Megakaryocytic Cell Lines: A Study Using a Novel Monoclonal Antibody against GPV

1994 ◽  
Vol 72 (05) ◽  
pp. 762-769 ◽  
Author(s):  
Toshiro Takafuta ◽  
Kingo Fujirmura ◽  
Hironori Kawano ◽  
Masaaki Noda ◽  
Tetsuro Fujimoto ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Cell Lines ◽  
Immunohistochemical Analysis ◽  
Specific Protein ◽  
Platelet Membrane ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Flow Cytometric ◽  
Polymerase Chain ◽  
Megakaryocytic Cell

SummaryGlycoprotein V (GPV) is a platelet membrane protein with a molecular weight of 82 kD, and one of the leucine rich glycoproteins (LRG). By reverse transcription-polymerase chain reaction (RT-PCR), GPV cDNA was amplified from mRNA of platelets and megakaryocytic cell lines. However, since there are few reports indicating whether GPV protein is expressed in megakaryocytes as a lineage and maturation specific protein, we studied the GPV expression at the protein level by using a novel monoclonal antibody (1D9) recognizing GPV. Flow cytometric and immunohistochemical analysis indicated that GPV was detected on the surface and in the cytoplasm of only the megakaryocytes in bone marrow aspirates. In a megakaryocytic cell line UT-7, GPV antigen increased after treatment with phorbol-12-myri-state-13-acetate (PMA). These data indicate that only megakaryocytes specifically express the GPV protein among hematopoietic cells and that the expression of GPV increases with differentiation of the megakaryocyte as GPIb-IX complex.

scholarly journals Super Magnetic Niosomal Nanocarrier as a New Approach for Treatment of Breast Cancer: A Case Study on SK-BR-3 and MDA-MB-231 Cell Lines

2021 ◽  
Vol 22 (15) ◽  
pp. 7948
Author(s):  
Elham Jamshidifar ◽  
Faten Eshrati Yeganeh ◽  
Mona Shayan ◽  
Mohammad Tavakkoli Yaraki ◽  
Mahsa Bourbour ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Cells ◽  
Cell Lines ◽  
Cancer Cell ◽  
Cellular Uptake ◽  
Targeted Delivery ◽  
Breast Cancer Cells ◽  
Cancer Cell Lines ◽  
Breast Cancer Cell Lines ◽  
Silica Layer

In the present study, a magnetic niosomal nanocarrier for co-delivery of curcumin and letrozole into breast cancer cells has been designed. The magnetic NiCoFe2O4 core was coated by a thin layer of silica, followed by a niosomal structure, allowing us to load letrozole and curcumin into the silica layer and niosomal layer, respectively, and investigate their synergic effects on breast cancer cells. Furthermore, the nanocarriers demonstrated a pH-dependent release due to the niosomal structure at their outer layer, which is a promising behavior for cancer treatment. Additionally, cellular assays revealed that the nanocarriers had low cellular uptake in the case of non-tumorigenic cells (i.e., MCF-10A) and related high viability but high cellular uptake in cancer cell lines (i.e., MDA-MB-231 and SK-BR-3) and related low viability, which is evidenced in their high cytotoxicity against different breast cancer cell lines. The cytotoxicity of the letrozole/curcumin co-loaded nanocarrier is higher than that of the aqueous solutions of both drugs, indicating their enhanced cellular uptake in their encapsulated states. In particular, NiCoFe2O4@L-Silica-L@C-Niosome showed the highest cytotoxicity effects on MDA-MB-231 and SK-BR-3 breast cancer cells. The observed cytotoxicity was due to regulation of the expression levels of the studied genes in breast cancer cells, where downregulation was observed for the Bcl-2, MMP 2, MMP 9, cyclin D, and cyclin E genes while upregulation of the expression of the Bax, caspase-3, and caspase-9 genes was observed. The flow cytometry results also revealed that NiCoFe2O4@L-Silica-L@C-Niosome enhanced the apoptosis rate in both MDA-MB-231 and SK-BR-3 cells compared to the control samples. The findings of our research show the potential of designing magnetic niosomal formulations for simultaneous targeted delivery of both hydrophobic and hydrophilic drugs into cancer cells in order to enhance their synergic chemotherapeutic effects. These results could open new avenues into the future of nanomedicine and the development of theranostic agents.

Cytometric Profiling of CD133+ Cells in Human Colon Carcinoma Cell Lines Identifies a Common core Phenotype and Cell Type-specific Mosaics

2013 ◽  
Vol 28 (3) ◽  
pp. 267-273 ◽  
Author(s):  
Marica Gemei ◽  
Rosa Di Noto ◽  
Peppino Mirabelli ◽  
Luigi Del Vecchio
Keyword(s):  
Colorectal Cancer ◽  
Stem Cells ◽  
Cancer Stem Cells ◽  
Colon Carcinoma ◽  
Cell Lines ◽  
Carcinoma Cell ◽  
Cell Type ◽  
Carcinoma Cell Lines ◽  
Cell Type Specific

In colorectal cancer, CD133+ cells from fresh biopsies proved to be more tumorigenic than their CD133– counterparts. Nevertheless, the function of CD133 protein in tumorigenic cells seems only marginal. Moreover, CD133 expression alone is insufficient to isolate true cancer stem cells, since only 1 out of 262 CD133+ cells actually displays stem-cell capacity. Thus, new markers for colorectal cancer stem cells are needed. Here, we show the extensive characterization of CD133+ cells in 5 different colon carcinoma continuous cell lines (HT29, HCT116, Caco2, GEO and LS174T), each representing a different maturation level of colorectal cancer cells. Markers associated with stemness, tumorigenesis and metastatic potential were selected. We identified 6 molecules consistently present on CD133+ cells: CD9, CD29, CD49b, CD59, CD151, and CD326. By contrast, CD24, CD26, CD54, CD66c, CD81, CD90, CD99, CD112, CD164, CD166, and CD200 showed a discontinuous behavior, which led us to identify cell type-specific surface antigen mosaics. Finally, some antigens, e.g. CD227, indicated the possibility of classifying the CD133+ cells into 2 subsets likely exhibiting specific features. This study reports, for the first time, an extended characterization of the CD133+ cells in colon carcinoma cell lines and provides a “dictionary” of antigens to be used in colorectal cancer research.

scholarly journals Susceptibility of Midge and Mosquito Vectors to SARS-CoV-2

2021 ◽  
Author(s):  
Velmurugan Balaraman ◽  
Barbara S Drolet ◽  
Natasha N Gaudreault ◽  
William C Wilson ◽  
Jeana Owens ◽  
...  
Keyword(s):  
Cell Lines ◽  
Potential Role ◽  
Animal Origin ◽  
Culex Tarsalis ◽  
Biting Midges ◽  
Bacterial Diseases ◽  
Culicoides Sonorensis ◽  
Insect Cell Lines ◽  
Intrathoracic Inoculation

Abstract SARS-CoV-2 is a recently emerged, highly contagious virus and the cause of the current COVID-19 pandemic. It is a zoonotic virus, although its animal origin is not clear yet. Person-to-person transmission occurs by inhalation of infected droplets and aerosols, or by direct contact with contaminated fomites. Arthropods transmit numerous viral, parasitic, and bacterial diseases; however, the potential role of arthropods in SARS-CoV-2 transmission is not fully understood. Thus far, a few studies have demonstrated that SARS-CoV-2 replication is not supported in cells from certain insect species nor in certain species of mosquitoes after intrathoracic inoculation. In this study, we expanded the work of SARS-CoV-2 susceptibility to biting insects after ingesting a SARS-CoV-2-infected bloodmeal. Species tested included Culicoides sonorensis (Wirth & Jones) (Diptera: Ceratopogonidae) biting midges, as well as Culex tarsalis (Coquillett) and Culex quinquefasciatus (Say) mosquitoes (Diptera: Culicidae), all known biological vectors for numerous RNA viruses. Arthropods were allowed to feed on SARS-CoV-2-spiked blood and at a time point postinfection analyzed for the presence of viral RNA and infectious virus. Additionally, cell lines derived from C. sonorensis (W8a), Aedes aegypti (Linnaeus) (Diptera: Culicidae) (C6/36), Cx. quinquefasciatus (HSU), and Cx. tarsalis (CxTrR2) were tested for SARS-CoV-2 susceptibility. Our results indicate that none of the biting insects, nor the insect cell lines evaluated support SARS-CoV-2 replication, suggesting that these species are unable to be biological vectors of SARS-CoV-2.

Analysis of the effect of DEK overexpression on the survival and proliferation of bone marrow stromal cells

2019 ◽  
Vol 44 (4) ◽  
pp. 510-516
Author(s):  
Türkan Çakar ◽  
Ayten Kandilci
Keyword(s):  
Cell Lines ◽  
Stromal Cells ◽  
Cell Sorting ◽  
Bone Marrow Stromal Cells ◽  
Nuclear Protein ◽  
Cell Type ◽  
Facs Analysis ◽  
Periodic Growth ◽  
Stromal Cell Line ◽  
Rpmi 8226

Abstract Objective DEK is ubiquitously expressed and encodes a nuclear protein, which is also released from some cells. Overexpression of DEK suppresses proliferation of some blood cell progenitors whereas it increases proliferation of epithelial tumors. We showed that DEK is overexpressed in BM cells of 12% of multiple myeloma (MM) patients. Here, we aimed to test if DEK overexpression effects the proliferation and viability of BM stromal cells or MM cells co-cultured with DEK-overexpressing stromal cells, mimicking the BM microenvironment. Methods DEK is stably overexpressed in the BM stromal cell line HS27A. Periodic growth curve and fluorescent activated cell sorting (FACS) analysis was performed to determine the effect of DEK overexpression on HS27A cells and MM cell lines (RPMI-8226 and U266) that are co-cultured with these HS27A cells. Results We showed that, on the contrary to blood progenitors or ephitelial cells, DEK overexpression doesn’t alter the viability or proliferation of the HS27A cells, or the MM cell lines which are co-cultured with DEK-overexpressing HS27A cells. Conclusions Our results suggest that effect of DEK overexpression on the proliferation is cell type and context dependent and increased DEK expression is tolerable by the stromal cells and the co-cultured MM cell lines without effecting proliferation and viability.

A sensitive flow cytometric method for measuring the oxidative burst

1997 ◽  
Vol 202 (2) ◽  
pp. 105-111 ◽  
Author(s):  
Michael A Model ◽  
Mark A KuKuruga ◽  
Robert F Todd
Keyword(s):  
Oxidative Burst ◽  
Flow Cytometric Method ◽  
Flow Cytometric
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