<i>De novo</i> identification of cell‐type specific antibody–antigen pairs by phage display subtraction

Stausbøl-Grøn, Brian; Jensen, Kim B.; Jensen, Kristian; Jensen, Michael Hansen; Clark, Brian F. C.

doi:10.1046/j.1432-1327.2001.02210.x

Cited by 8 publications

(6 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Phage display is emerging as a promising and useful technique in the expanding field of proteomics (21,38,55), although it is still in its infancy in this respect. The postgenomic era of molecular and cellular biology calls for methods that are capable of analyzing and identifying differentially expressed proteins to analyze the complexity of the functional genome.…”

Section: Fig 3 Immunoprecipitation Ofmentioning

confidence: 99%

Identification of Keratinocyte-specific Markers Using Phage Display and Mass Spectrometry

Jensen

Ravn

et al. 2003

Molecular & Cellular Proteomics

Self Cite

View full text Add to dashboard Cite

Specific molecular markers for various normal and pathogenic cell states and cell types provide knowledge of basic biological systems and have a direct application in targeted therapy. We describe a proteomic method based on the combination of new and improved phage display antibody technologies and mass spectrometry that allows identification of cell type-specific protein markers. The most important features of the method are (i) reduction of experimental noise originating from background binding of phage particles and (ii) isolation of affinity binders after a single round of selection, which assures a high diversity of binders. The method demonstrates, for the first time, the ability to detect, identify, and analyze both secreted and membrane-associated extracellular proteins as well as a variety of different cellular structures including proteins and carbohydrates. The optimized phage display method was applied to analysis of human skin keratinocytes resulting in the isolation of a panel of antibodies. Fourteen of these antibodies were further characterized, half of which predominantly recognized keratinocytes in a screen of a range of different cell types. Three cognate keratinocyte antigens were subsequently identified by mass spectrometry as laminin-5, plectin, and fibronectin. The combination of phage display technology with mass spectrometry methods for protein identification is a general and promising approach for proteomic analysis of cell surface complexity.

show abstract

Section: Fig 3 Immunoprecipitation Ofmentioning

confidence: 99%

Identification of Keratinocyte-specific Markers Using Phage Display and Mass Spectrometry

Jensen

Ravn

et al. 2003

Molecular & Cellular Proteomics

Self Cite

View full text Add to dashboard Cite

show abstract

“…Furthermore, mRNA levels are not informative of proteins that are post-translationally modified. Screening phage display libraries offers the unbiased evaluation of all proteins whose expression levels change in response to carcinogen exposure (11). While phage display-based screening of the complete array of cellular proteins does not guarantee detection of causality, this unbiased screening approach, in general, is a significant step forward from the traditional arbitrary selection of a few proteins that may or may not be causally related to tumorigenesis.…”

Section: Introductionmentioning

confidence: 99%

Carcinogen-induced histone alteration in normal human mammary epithelial cells

Bradley

Meer²,

Roodi³

et al. 2007

Carcinogenesis

View full text Add to dashboard Cite

Little is known about early carcinogen-induced protein alterations in mammary epithelium. Detection of early alterations would enhance our understanding of early-stage carcinogenesis. Here, normal human mammary epithelial cells (HMECs) were exposed to dietary and environmental carcinogens [2-amino-1-methyl-6-phenylimidazo[4,5b]pyridine (PhIP), 4-aminobiphenyl (ABP), benzo[a]pyrene, 2,3,7,8-tetrachlorodibenzo-p-dioxin] individually or in combination. A phage display library of single-chain variable fragment antibodies was used to screen protein targets altered by the treatment. In combination with matrix-assisted laser desorption time of flight, we identified histone H3 as a target antigen. Although histone H3 total protein remained unchanged in control and treated HMEC, the methylation of lysine 4 was altered. A reduction in mono-methyl histone H3 (Lys 4) was observed in treated HMEC compared with control HMEC. This alteration was shown to be dependent on carcinogen concentration and specific for PhIP and ABP. To characterize potential histone demethylation mechanisms, localization and protein expression patterns of lysine-specific demethylase 1 (LSD1) were analyzed. In control HMEC, LSD1 was present at the nuclear periphery. However, following 72 h carcinogen treatment, LSD1 localized within the nucleus. Within 48 h after treatment, mono-methyl histone H3 (Lys 4) was restored and LSD1 localization was reversed. Protein expression levels of LSD1 were also increased in treated HMEC compared with control HMEC. Our data suggest that the induction of a single enzyme, LSD1, represents an early response to carcinogen exposure, which leads to the demethylation of histone H3 (Lys 4), which, in turn, may influence the expression of multiple genes critical in early-stage mammary carcinogenesis.

show abstract

“…Since in vitro differentiation does not lead to fully differentiated corneocytes, four early, suprabasal markers were chosen: cytokeratins, K10 (ab9025, mouse monoclonal; Abcam, Cambridge, UK) and K14 (as single-chain variable fragments [scFv] 11 ), and involucrin (ab14504, mouse monoclonal; Abcam Cambridge, UK). In addition, the scFvs clone 10, which is specific for keratinocytes, but whose antigen is not known, was also tested.…”

Section: Differentiation Markers and Elisamentioning

confidence: 99%

Kinetin‐Induced Differentiation of Normal Human Keratinocytes Undergoing Aging in Vitro

Berge

2006

Annals of the New York Academy of Sciences

View full text Add to dashboard Cite

Kinetin (N(6)-furfuryladenine) is a cytokinin growth factor having several anti-aging effects reported for human cells and fruit flies. We have observed that short-term culturing of human keratinocytes in the presence of 40 to 200 microM kinetin results in a significant inhibition of cell growth. Studies were undertaken to analyze the process of differentiation as a reason for growth inhibition. Keratinocytes at different passage levels were treated with fetal calf serum (FCS) and calcium as differentiation-inducing positive controls, with different concentrations of kinetin, and with a combination of kinetin and calcium. The induction and progression of differentiation was monitored by morphological observations and by using several differentiation markers, including keratins (K10 and K14), involucrin, epidermal transglutaminase, and some new keratinocyte-specific antibodies isolated by the phage display method. In young keratinocytes, two days of calcium treatment reduced the K14 level by 78%, and increased the levels of K10 and involucrin by 40% and 29%, respectively. In comparison, 40 microM kinetin had no effect on the K14 level, but increased the K10 level by 28% and that of involucrin by four-fold. The combination of calcium and 40 microM kinetin led to a decrease by 23% in the K14 level, to an increase in the level of K10 by 55%, and to a two-fold rise in the involucrin level. These results suggest that the rate, extent, and quality of differentiation depend on the inducing agent, and that kinetin may be useful in promoting the differentiation of human keratinocytes, especially in the presence of calcium.

show abstract

De novo identification of cell‐type specific antibody–antigen pairs by phage display subtraction

Cited by 8 publications

References 30 publications

Identification of Keratinocyte-specific Markers Using Phage Display and Mass Spectrometry

Identification of Keratinocyte-specific Markers Using Phage Display and Mass Spectrometry

Carcinogen-induced histone alteration in normal human mammary epithelial cells

Kinetin‐Induced Differentiation of Normal Human Keratinocytes Undergoing Aging in Vitro

Contact Info

Product

Resources

About