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Article: Circulating melanoma cells: scoping the target.

Joshi, Powrnima / Zborowski, Maciej / Triozzi, Pierre L

2013 Volume 3, Page(s) 189

Language	English
Publishing date	2013-08-12
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2649216-7
ISSN	2234-943X
ISSN	2234-943X
DOI	10.3389/fonc.2013.00189
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Article ; Online: Microfluidic chip for graduated magnetic separation of circulating tumor cells by their epithelial cell adhesion molecule expression and magnetic nanoparticle binding.

Williams, P Stephen / Moore, Lee R / Joshi, Powrnima / Goodin, Mark / Zborowski, Maciej / Fleischman, Aaron

Journal of chromatography. A

2020 Volume 1637, Page(s) 461823

Abstract: The enumeration of circulating tumor cells (CTCs) in the peripheral bloodstream of metastatic cancer patients has contributed to improvements in prognosis and therapeutics. There have been numerous approaches to capture and counting of CTCs. However, ... ...

Abstract	The enumeration of circulating tumor cells (CTCs) in the peripheral bloodstream of metastatic cancer patients has contributed to improvements in prognosis and therapeutics. There have been numerous approaches to capture and counting of CTCs. However, CTCs have potential information beyond simple enumeration and hold promise as a liquid biopsy for cancer and a pathway for personalized cancer therapy by detecting the subset of CTCs having the highest metastatic potential. There is evidence that epithelial cell adhesion molecule (EpCAM) expression level distinguishes these highly metastatic CTCs. The few previous approaches to selective CTC capture according to EpCAM expression level are reviewed. A new two-stage microfluidic device for separation, enrichment and release of CTCs into subpopulations sorted by EpCAM expression level is presented here. It relies upon immunospecific magnetic nanoparticle labeling of CTCs followed by their field- and flow-based separation in the first stage and capture as discrete subpopulations in the second stage. To fine tune the separation, the magnetic field profile across the first stage microfluidic channel may be modified by bonding small Vanadium Permendur strips to its outer walls. Mathematical modeling of magnetic fields and fluid flows supports the soundness of the design.
MeSH term(s)	Cell Line, Tumor ; Cell Separation/instrumentation ; Epithelial Cell Adhesion Molecule/metabolism ; Humans ; Lab-On-A-Chip Devices ; Magnetics/instrumentation ; Neoplastic Cells, Circulating ; Oligonucleotide Array Sequence Analysis ; Protein Binding
Chemical Substances	Epithelial Cell Adhesion Molecule
Language	English
Publishing date	2020-12-17
Publishing country	Netherlands
Document type	Journal Article
ZDB-ID	1171488-8
ISSN	1873-3778 ; 0021-9673
ISSN (online)	1873-3778
ISSN	0021-9673
DOI	10.1016/j.chroma.2020.461823
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Article ; Online: Gene-gun transfection of hippocampal neurons.

Joshi, Powrnima / Dunaevsky, Anna

Journal of visualized experiments : JoVE

2006 , Issue 1, Page(s) 121

MeSH term(s)	Biolistics ; Hippocampus/cytology ; Hippocampus/metabolism ; Neurons/metabolism ; Transfection/methods
Language	English
Publishing date	2006-11-30
Publishing country	United States
Document type	Journal Article ; Video-Audio Media
ZDB-ID	2259946-0
ISSN	1940-087X ; 1940-087X
ISSN (online)	1940-087X
ISSN	1940-087X
DOI	10.3791/121
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Article ; Online: Expression of natural killer cell regulatory microRNA by uveal melanoma cancer stem cells.

Joshi, Powrnima / Kooshki, Mitra / Aldrich, Wayne / Varghai, Daniel / Zborowski, Maciej / Singh, Arun D / Triozzi, Pierre L

Clinical & experimental metastasis

2016 Volume 33, Issue 8, Page(s) 829–838

Abstract: Natural killer (NK) cells are implicated in the control of metastasis in uveal melanoma, a process that has been ascribed to its cancer stem cell subpopulation. NK cell activation is regulated by specific microRNA (miR). The NK cell sensitivity and ... ...

Abstract	Natural killer (NK) cells are implicated in the control of metastasis in uveal melanoma, a process that has been ascribed to its cancer stem cell subpopulation. NK cell activation is regulated by specific microRNA (miR). The NK cell sensitivity and regulatory miR production of uveal melanoma cancer stem cells was examined. Cancer stem cells enriched from aggressively metastatic MUM2B uveal melanoma cells by selecting CD271
MeSH term(s)	Adapalene/immunology ; Adapalene/metabolism ; Cytotoxicity, Immunologic/immunology ; Gene Expression Regulation, Neoplastic ; Humans ; Killer Cells, Natural/metabolism ; Killer Cells, Natural/pathology ; Melanocytes/pathology ; Melanoma/genetics ; Melanoma/pathology ; MicroRNAs/biosynthesis ; MicroRNAs/genetics ; Neoplastic Stem Cells/metabolism ; Neoplastic Stem Cells/pathology ; Transfection ; Uveal Neoplasms/genetics ; Uveal Neoplasms/pathology
Chemical Substances	MIRN155 microRNA, human ; MicroRNAs ; Adapalene (1L4806J2QF)
Language	English
Publishing date	2016-08-26
Publishing country	Netherlands
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	604952-7
ISSN	1573-7276 ; 0262-0898
ISSN (online)	1573-7276
ISSN	0262-0898
DOI	10.1007/s10585-016-9815-9
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Article ; Online: A quantitative determination of magnetic nanoparticle separation using on-off field operation of quadrupole magnetic field-flow fractionation (QMgFFF).

Orita, Toru / Moore, Lee R / Joshi, Powrnima / Tomita, Masahiro / Horiuchi, Takashi / Zborowski, Maciej

Analytical sciences : the international journal of the Japan Society for Analytical Chemistry

2013 Volume 29, Issue 7, Page(s) 761–764

Abstract: Quadrupole Magnetic Field-Flow Fractionation (QMgFFF) is a technique for characterization of sub-micrometer magnetic particles based on their retention in the magnetic field from flowing suspensions. Different magnetic field strengths and volumetric flow ...

Abstract	Quadrupole Magnetic Field-Flow Fractionation (QMgFFF) is a technique for characterization of sub-micrometer magnetic particles based on their retention in the magnetic field from flowing suspensions. Different magnetic field strengths and volumetric flow rates were tested using on-off field application and two commercial nanoparticle preparations that significantly differed in their retention parameter, λ (by nearly 8-fold). The fractograms showed a regular pattern of higher retention (98.6% v. 53.3%) for the larger particle (200 nm v. 90 nm) at the higher flow rate (0.05 mL/min v. 0.01 mL/min) at the highest magnetic field (0.52 T), as expected because of its lower retention parameter. The significance of this approach is a demonstration of a system that is simpler in operation than a programmed field QMgFFF in applications to particle mixtures consisting of two distinct particle fractions. This approach could be useful for detection of unwanted particulate contaminants, especially important in industrial and biomedical applications.
MeSH term(s)	Fractionation, Field Flow/methods ; Magnetics ; Nanoparticles
Language	English
Publishing date	2013-07-10
Publishing country	Switzerland
Document type	Journal Article ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	1483376-1
ISSN	1348-2246 ; 1348-2246
ISSN (online)	1348-2246
ISSN	1348-2246
DOI	10.2116/analsci.29.761
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Article ; Online: Hair cell BK channels interact with RACK1, and PKC increases its expression on the cell surface by indirect phosphorylation.

Surguchev, Alexei / Bai, Jun-Ping / Joshi, Powrnima / Navaratnam, Dhasakumar

American journal of physiology. Cell physiology

2012 Volume 303, Issue 2, Page(s) C143–50

Abstract: Large conductance (BK) calcium activated potassium channels (Slo) are ubiquitous and implicated in a number of human diseases including hypertension and epilepsy. BK channels consist of a pore forming α-subunit (Slo) and a number of accessory subunits. ... ...

Abstract	Large conductance (BK) calcium activated potassium channels (Slo) are ubiquitous and implicated in a number of human diseases including hypertension and epilepsy. BK channels consist of a pore forming α-subunit (Slo) and a number of accessory subunits. In hair cells of nonmammalian vertebrates these channels play a critical role in electrical resonance, a mechanism of frequency selectivity. Hair cell BK channel clusters on the surface and currents increase along the tonotopic axis and contribute significantly to the responsiveness of these hair cells to sounds of high frequency. In contrast, messenger RNA levels encoding the Slo gene show an opposite decrease in high frequency hair cells. To understand the molecular events underlying this paradox, we used a yeast two-hybrid screen to isolate binding partners of Slo. We identified Rack1 as a Slo binding partner and demonstrate that PKC activation increases Slo surface expression. We also establish that increased Slo recycling of endocytosed Slo is at least partially responsible for the increased surface expression of Slo. Moreover, analysis of several PKC phosphorylation site mutants confirms that the effects of PKC on Slo surface expression are likely indirect. Finally, we show that Slo clusters on the surface of hair cells are also increased by increased PKC activity and may contribute to the increasing amounts of channel clusters on the surface of high-frequency hair cells.
MeSH term(s)	Animals ; Cell Membrane/genetics ; Cell Membrane/metabolism ; Chickens ; Cochlea/metabolism ; GTP-Binding Proteins/biosynthesis ; GTP-Binding Proteins/genetics ; GTP-Binding Proteins/metabolism ; Gene Expression Regulation ; HEK293 Cells ; Hair Cells, Auditory/metabolism ; Hair Cells, Auditory/physiology ; Humans ; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits/genetics ; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits/metabolism ; Large-Conductance Calcium-Activated Potassium Channels/genetics ; Large-Conductance Calcium-Activated Potassium Channels/metabolism ; Neoplasm Proteins/biosynthesis ; Neoplasm Proteins/genetics ; Neoplasm Proteins/metabolism ; Phosphorylation/genetics ; Protein Kinase C/biosynthesis ; Protein Kinase C/genetics ; Protein Kinase C/physiology ; Receptors for Activated C Kinase ; Receptors, Cell Surface/biosynthesis ; Receptors, Cell Surface/genetics ; Receptors, Cell Surface/metabolism ; Up-Regulation/genetics
Chemical Substances	KCNMA1 protein, human ; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits ; Large-Conductance Calcium-Activated Potassium Channels ; Neoplasm Proteins ; RACK1 protein, human ; Receptors for Activated C Kinase ; Receptors, Cell Surface ; Protein Kinase C (EC 2.7.11.13) ; GTP-Binding Proteins (EC 3.6.1.-)
Language	English
Publishing date	2012-04-25
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	392098-7
ISSN	1522-1563 ; 0363-6143
ISSN (online)	1522-1563
ISSN	0363-6143
DOI	10.1152/ajpcell.00062.2012
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Article: Circular Halbach Array for Fast Magnetic Separation of Hyaluronan-Expressing Tissue Progenitors

Joshi, Powrnima / Boehm Cynthia / Caralla Tonya / Moore Lee R / Muschler George / Williams P. Stephen / Zborowski Maciej

Analytical chemistry. 2015 Oct. 06, v. 87, no. 19

2015

Abstract: Connective tissue progenitors (CTPs) are a promising therapeutic agent for bone repair. Hyaluronan, a high molecular mass glycosaminoglycan, has been shown by us to be a suitable biomarker for magnetic separation of CTPs from bone marrow aspirates in a ... ...

Abstract	Connective tissue progenitors (CTPs) are a promising therapeutic agent for bone repair. Hyaluronan, a high molecular mass glycosaminoglycan, has been shown by us to be a suitable biomarker for magnetic separation of CTPs from bone marrow aspirates in a canine model. For the therapy to be applicable in humans, the magnetic separation process requires scale-up without compromising the viability of the cells. The scaled-up device presented here utilizes a circular Halbach array of diametrically magnetized, cylindrical permanent magnets. This allows precise control of the magnetic field gradient driving the separation, with theoretical analysis favoring a hexapole field. The separation vessel has the external diameter of a 50 mL conical centrifuge tube and has an internal rod that excludes cells from around the central axis. The magnet and separation vessel (collectively dubbed the hexapole magnet separator or HMS) was tested on four human and four canine bone marrow aspirates. Each CTP-enriched cell product was tested using cell culture bioassays as surrogates for in vivo engraftment quality. The magnetically enriched cell fractions showed statistically significant, superior performance compared to the unenriched and depleted cell fractions for all parameters tested, including CTP prevalence (CTPs per 10â¶ nucleated cells), proliferation by colony forming unit (CFU) counts, and differentiation by staining for the presence of osteogenic and chondrogenic cells. The simplicity and speed of the HMS operation could allow both CTP isolation and engraftment during a single surgical procedure, minimizing trauma to patients and lowering cost to health care providers.
Keywords	bioassays ; biomarkers ; bone marrow ; cell culture ; cell viability ; dogs ; health care costs ; health care workers ; humans ; hyaluronic acid ; magnetic fields ; magnetic materials ; magnetic separation ; models ; molecular weight ; patients ; staining ; surgery
Language	English
Dates of publication	2015-1006
Size	p. 9908-9915.
Publishing place	American Chemical Society
Document type	Article
ZDB-ID	1508-8
ISSN	1520-6882 ; 0003-2700
ISSN (online)	1520-6882
ISSN	0003-2700
DOI	10.1021%2Facs.analchem.5b02431
Database	NAL-Catalogue (AGRICOLA)

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Article ; Online: Circular Halbach array for fast magnetic separation of hyaluronan-expressing tissue progenitors.

Joshi, Powrnima / Williams, P Stephen / Moore, Lee R / Caralla, Tonya / Boehm, Cynthia / Muschler, George / Zborowski, Maciej

Analytical chemistry

2015 Volume 87, Issue 19, Page(s) 9908–9915

Abstract	Connective tissue progenitors (CTPs) are a promising therapeutic agent for bone repair. Hyaluronan, a high molecular mass glycosaminoglycan, has been shown by us to be a suitable biomarker for magnetic separation of CTPs from bone marrow aspirates in a canine model. For the therapy to be applicable in humans, the magnetic separation process requires scale-up without compromising the viability of the cells. The scaled-up device presented here utilizes a circular Halbach array of diametrically magnetized, cylindrical permanent magnets. This allows precise control of the magnetic field gradient driving the separation, with theoretical analysis favoring a hexapole field. The separation vessel has the external diameter of a 50 mL conical centrifuge tube and has an internal rod that excludes cells from around the central axis. The magnet and separation vessel (collectively dubbed the hexapole magnet separator or HMS) was tested on four human and four canine bone marrow aspirates. Each CTP-enriched cell product was tested using cell culture bioassays as surrogates for in vivo engraftment quality. The magnetically enriched cell fractions showed statistically significant, superior performance compared to the unenriched and depleted cell fractions for all parameters tested, including CTP prevalence (CTPs per 10(6) nucleated cells), proliferation by colony forming unit (CFU) counts, and differentiation by staining for the presence of osteogenic and chondrogenic cells. The simplicity and speed of the HMS operation could allow both CTP isolation and engraftment during a single surgical procedure, minimizing trauma to patients and lowering cost to health care providers.
MeSH term(s)	Animals ; Bone Marrow Cells/cytology ; Cell Differentiation ; Cell Separation/instrumentation ; Cells, Cultured ; Dogs ; Equipment Design ; Humans ; Hyaluronic Acid/analysis ; Magnetics/instrumentation ; Stem Cells/cytology
Chemical Substances	Hyaluronic Acid (9004-61-9)
Language	English
Publishing date	2015-09-14
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, U.S. Gov't, Non-P.H.S.
ZDB-ID	1508-8
ISSN	1520-6882 ; 0003-2700
ISSN (online)	1520-6882
ISSN	0003-2700
DOI	10.1021/acs.analchem.5b02431
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Article ; Online: Enrichment of circulating melanoma cells (CMCs) using negative selection from patients with metastatic melanoma.

Joshi, Powrnima / Jacobs, Barbara / Derakhshan, Adeeb / Moore, Lee R / Elson, Paul / Triozzi, Pierre L / Borden, Ernest / Zborowski, Maciej

Oncotarget

2014 Volume 5, Issue 9, Page(s) 2450–2461

Abstract: Circulating tumor cells have emerged as prognostic biomarkers in the treatment of metastatic cancers of epithelial origins viz., breast, colorectal and prostate. These tumors express Epithelial Cell Adhesion Molecule (EpCAM) on their cell surface which ... ...

Abstract	Circulating tumor cells have emerged as prognostic biomarkers in the treatment of metastatic cancers of epithelial origins viz., breast, colorectal and prostate. These tumors express Epithelial Cell Adhesion Molecule (EpCAM) on their cell surface which is used as an antigen for immunoaffinity capture. However, EpCAM capture technologies are of limited utility for non-epithelial cancers such as melanoma. We report a method to enrich Circulating Melanoma Cells (CMCs) that does not presuppose malignant cell characteristics. CMCs were enriched by centrifugation of blood samples from healthy (N = 10) and patient (N = 11) donors, followed by RBC lysis and immunomagnetic depletion of CD45-positive leukocytes in a specialized magnetic separator. CMCs were identified by immunocytochemistry using Melan-A or S100B as melanoma markers and enumerated using automated microscopy image analyses. Separation was optimized for maximum sensitivity and recovery of CMCs. Our results indicate large number of CMCs in Stage IV melanoma patients. Analysis of survival suggested a trend toward decreased survival with increased number of CMCs. Moreover, melanoma-associated miRs were found to be higher in CMC-enriched fractions in two patients when compared with the unseparated samples, validating this method as applicable for molecular analyses. Negative selection is a promising approach for isolation of CMCs and other EpCAM -negative CTCs, and is amenable to molecular analysis of CMCs. Further studies are required to validate its efficacy at capturing specific circulating cells for genomic analysis, and xenograft studies.
MeSH term(s)	Biomarkers, Tumor/analysis ; Case-Control Studies ; Cell Separation ; Humans ; Immunoenzyme Techniques ; Immunomagnetic Separation/methods ; Leukocyte Common Antigens/blood ; MART-1 Antigen/blood ; Melanoma/blood ; MicroRNAs/genetics ; Neoplastic Cells, Circulating/chemistry ; Neoplastic Cells, Circulating/pathology ; Prognosis ; RNA, Messenger/genetics ; Real-Time Polymerase Chain Reaction ; Reverse Transcriptase Polymerase Chain Reaction ; S100 Calcium Binding Protein beta Subunit/blood ; Survival Rate ; Tumor Cells, Cultured
Chemical Substances	Biomarkers, Tumor ; MART-1 Antigen ; MicroRNAs ; RNA, Messenger ; S100 Calcium Binding Protein beta Subunit ; S100B protein, human ; Leukocyte Common Antigens (EC 3.1.3.48) ; PTPRC protein, human (EC 3.1.3.48)
Language	English
Publishing date	2014-05-10
Publishing country	United States
Document type	Journal Article
ZDB-ID	2560162-3
ISSN	1949-2553 ; 1949-2553
ISSN (online)	1949-2553
ISSN	1949-2553
DOI	10.18632/oncotarget.1683
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Article ; Online: CDK5 interacts with Slo and affects its surface expression and kinetics through direct phosphorylation.

Bai, Jun-Ping / Surguchev, Alexei / Joshi, Powrnima / Gross, Liza / Navaratnam, Dhasakumar

American journal of physiology. Cell physiology

2011 Volume 302, Issue 5, Page(s) C766–80

Abstract: Large-conductance calcium-activated potassium (BK) channels are ubiquitous and play an important role in a number of diseases. In hair cells of the ear, they play a critical role in electrical tuning, a mechanism of frequency discrimination. These ... ...

Abstract	Large-conductance calcium-activated potassium (BK) channels are ubiquitous and play an important role in a number of diseases. In hair cells of the ear, they play a critical role in electrical tuning, a mechanism of frequency discrimination. These channels show variable kinetics and expression along the tonotopic axis. Although the molecular underpinnings to its function in hair cells are poorly understood, it is established that BK channels consist of a pore-forming α-subunit (Slo) and a number of accessory subunits. Here we identify CDK5, a member of the cyclin-dependent kinase family, as an interacting partner of Slo. We show CDK5 to be present in hair cells and expressed in high concentrations in the cuticular plate and in the circumferential zone. In human embryonic kidney cells, we show that CDK5 inhibits surface expression of Slo by direct phosphorylation of Slo. Similarly, we note that CDK5 affects Slo voltage activation and deactivation kinetics, by a direct phosphorylation of T847. Taken together with its increasing expression along the tonotopic axis, these data suggest that CDK5 likely plays a critical role in electrical tuning and surface expression of Slo in hair cells.
MeSH term(s)	Animals ; Calcium Signaling/physiology ; Cell Membrane/genetics ; Cell Membrane/metabolism ; Chickens ; Cochlea/metabolism ; Cyclin-Dependent Kinase 5/metabolism ; Fluorescence Resonance Energy Transfer ; Gene Library ; HEK293 Cells ; Hair Cells, Auditory/metabolism ; Humans ; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits/genetics ; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits/metabolism ; Membrane Potentials/physiology ; Patch-Clamp Techniques ; Phosphorylation ; Two-Hybrid System Techniques ; Xenopus laevis
Chemical Substances	Large-Conductance Calcium-Activated Potassium Channel alpha Subunits ; Cyclin-Dependent Kinase 5 (EC 2.7.11.1)
Language	English
Publishing date	2011-11-16
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	392098-7
ISSN	1522-1563 ; 0363-6143
ISSN (online)	1522-1563
ISSN	0363-6143
DOI	10.1152/ajpcell.00339.2011
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