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Dive into the research topics where Miranda Yap is active.

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Featured researches published by Miranda Yap.


Stem Cells | 2008

Selection against undifferentiated human embryonic stem cells by a cytotoxic antibody recognizing podocalyxin-like protein-1.

Heng Liang Tan; Sheu Ngo Ang; Wey Jia Fong; Angela Chin; Jennifer Lo; Lu Zheng; Hannes Hentze; Robin Philp; Steve Oh; Miranda Yap

Future therapeutic applications of differentiated human embryonic stem cells (hESC) carry a risk of teratoma formation by contaminating undifferentiated hESC. We generated 10 monoclonal antibodies (mAbs) against surface antigens of undifferentiated hESC, showing strong reactivity against undifferentiated, but not differentiated hESC. The mAbs did not cross react with mouse fibroblasts and showed weak to no reactivity against human embryonal carcinoma cells. Notably, one antibody (mAb 84) is cytotoxic to undifferentiated hESC and NCCIT cells in a concentration‐dependent, complement‐independent manner. mAb 84 induced cell death of undifferentiated, but not differentiated hESC within 30 minutes of incubation, and immunoprecipitation of the mAb‐antigen complex revealed that the antigen is podocalyxin‐like protein‐1. Importantly, we observed absence of tumor formation when hESC and NCCIT cells were treated with mAb 84 prior to transplantation into severe combined immunodeficiency mice. Our data indicate that mAb 84 may be useful in eliminating residual hESC from differentiated cells populations for clinical applications.


Stem Cells | 2009

mAb 84, a Cytotoxic Antibody that Kills Undifferentiated Human Embryonic Stem Cells via Oncosis

Heng Liang Tan; Wey Jia Fong; Eng Hin Lee; Miranda Yap

The monoclonal antibody mAb 84, which binds to podocalyxin‐like protein‐1 (PODXL) on human embryonic stem cells (hESCs), was previously reported to bind and kill undifferentiated cells in in vitro and in vivo assays. In this study, we investigate the mechanism responsible for mAb 84‐induced hESCs cytotoxicity. Apoptosis was likely not the cause of mAb 84‐mediated cell death because no elevation of caspase activities or increased DNA fragmentation was observed in hESCs following incubation with mAb 84. Instead, it was preceded by cell aggregation and damage to cell membranes, resulting in the uptake of propidium iodide, and the leakage of intracellular sodium ions. Furthermore, examination of the cell surface by scanning electron microscopy revealed the presence of pores on the cell surface of mAb 84‐treated cells, which was absent from the isotype control. This mechanism of cell death resembles that described for oncosis, a form of cell death resulting from membrane damage. Additional data suggest that the binding of mAb 84 to hESCs initiates a sequence of events prior to membrane damage, consistent with oncosis. Degradation of actin‐associated proteins, namely, α‐actinin, paxillin, and talin, was observed. The perturbation of these actin‐associated proteins consequently permits the aggregation of PODXL, thus leading to the formation of pores. To our knowledge, this is the first report of oncotic cell death with hESCs as a model. STEM CELLS 2009;27:1792–1801


Biotechnology Advances | 2009

Developing genomic platforms for Chinese hamster ovary cells

Anne Kantardjieff; Peter Morin Nissom; Song Hui Chuah; Faraaz Noor Khan Yusufi; Nitya M. Jacob; Bhanu Chandra Mulukutla; Miranda Yap; Wei Shou Hu

Chinese hamster ovary (CHO) cells are widely used in recombinant protein production, yet despite their importance in bioprocessing, few genomic resources have been developed for this cell line. Over the past several years, we have made considerable progress in the development of genomic tools for CHO. Using Sanger-based sequencing technology, we have accrued a sequence repertoire of more than 68,000 expressed sequence tags (ESTs), representing more than 28,000 unique CHO transcripts. Using closely related species, we have functionally annotated this sequence set and have currently achieved significant representation in a number of functional classes, including some closely tied to recombinant protein production. This sequence repository has been used to design custom CHO Affymetrix arrays for transcriptome analysis. Illumina Solexa deep sequencing technology was also applied to study the CHO cell transcriptome and survey the identity and expression of small RNAs. These applications demonstrate the utility of genomic tools, and illustrate the applicability of emerging next-generation sequencing technologies.


Biotechnology and Bioengineering | 2011

Conserved microRNAs in Chinese hamster ovary cell lines.

Kathryn C. Johnson; Nitya M. Jacob; Peter Morin Nissom; Matthias Hackl; Lim Hseuh Lee; Miranda Yap; Wei Shou Hu

MicroRNAs (miRNAs), a class of short (20-24 nt) non-coding RNAs that direct post-transcriptional repression of messenger RNAs, increasingly have been shown to play a key role in regulating cellular physiology. We investigated the prevalence of miRNAs in Chinese hamster ovary (CHO) cells by high-throughput sequencing. Six cDNA libraries of small RNAs from four CHO cell lines were constructed and sequenced by Illumina sequencing. Three hundred fifty distinct miRNA and miRNA* sequences were identified through homology with other species, including mouse, rat, and human. While the majority of the identified miRNAs appear to be expressed ubiquitously, many miRNAs were found to have a wide range of expression levels between cell lines. The identification of these miRNAs will facilitate investigations of their contribution to the hyperproductivity trait.


Biotechnology and Bioengineering | 2010

Reaching the depth of the Chinese hamster ovary cell transcriptome

Nitya M. Jacob; Anne Kantardjieff; Faraaz Noor Khan Yusufi; Ernest F. Retzel; Bhanu Chandra Mulukutla; Song Hui Chuah; Miranda Yap; Wei Shou Hu

The high-throughput DNA sequencing Illumina Solexa GAII platform was employed to characterize the transcriptome of an antibody-producing Chinese hamster ovary (CHO) cell line. More than 55 million sequencing reads were generated and mapped to an existing set of CHO unigenes derived from expressed sequence tags (ESTs), as well as several public sequence databases. A very significant fraction of sequencing reads has not been previously seen. The frequency with which fragments of a unigene were sequenced was taken as an estimate of the abundance level of the corresponding transcripts. A wide dynamic range of transcript abundance levels was observed, spanning six orders of magnitude. However, the distribution of coverage across transcript lengths was found to vary, from relatively uniform to highly variable. This observation suggests that more challenges are yet to be resolved before direct sequencing can be used as a true quantitative measure of transcript level and for differential gene expression analysis. With the depth that high-throughput sequencing methods can reach, one can expect that the entire transcriptome of this industrially important organism will be decoded in the near future.


Journal of Immunological Methods | 1994

Enhanced IgG production in eRDF media with and without serum: A comparative study

Florence Chua; Steve K.W. Oh; Miranda Yap; W.K. Teo

The performance of three basal media RPMI, DMEM/F12 (DF) and eRDF (enhanced RDF, RPMI:DMEM:F12 in 2:1:1) were evaluated in cultures with and without serum with respect to cell proliferation, metabolism and monoclonal antibody (Mab) productivity. Based on the ease of adaptation, growth rate, maximum cell density and Mab production, the media were ranked as follows: eRDF > DF > RPMI. This was true for serum-free (SF) and serum supplemented (SS) media in static and shaker cultures. Growth performances in static and shaker cultures were consistently 20-50% lower in all three SF media compared to the corresponding SS conditions. Antibody titres in DF/SF and RPMI/SF cultures, irrespective of the culture condition, were generally similar or slightly lower than their SS counterparts. However, eRDF/SF medium yielded a much higher Mab titre (193 mg l-1) compared to eRDF/SS medium (145 mg l-1). This was also six times higher than the lowest titre of 30 mg l-1 in RPMI/SF medium. Hybridomas in eRDF/SF were further adapted to media without bovine serum albumin (eRDF/SF-BSA). Maximum cell densities in these cultures improved with scale up, from 1.1 x 10(6) ml-1 in static, to 1.9 x 10(6) ml-1 in shaker flasks, to 2.5 x 10(6) ml-1 in bioreactors. However, Ig levels remained between 100-130 mg l-1 which were much lower than in eRDF/SF medium. Thus BSA appears to be necessary for Ig production. The manufacturing cost (excluding purification) of Ig using eRDF was calculated to be between 17-50% of the price of the other two media and therefore this is regarded as the best medium for Ig production.


Archive | 2004

Genomic exploration on Chinese hamster ovary cells

M. De Leon Gatti; Katie F. Wlaschin; Anette Rink; A. Sanny; Kher Shing Tan; Peter Morin Nissom; Peh Fern Ong; Kathy Wong; R.J. Philip; B. Cham; C.F. Wong; K.M. Lim; Miranda Yap; Wei Shou Hu

Industrially important recombinant CHO cell clones carry many desirable phenotypic traits that differentiate them from the parental clone. The molecular basis for these traits at the genomic level is not well understood. A better understanding of key genetic alterations related to these traits will greatly facilitate the development of new cell lines and new processes. To advance this goal, we are constructing a CHO cDNA microarray. A phage library was constructed using mRNA pooled from different CHO clones under different cultivation conditions. Over four thousand sequences have been randomly isolated and sequenced. A cDNA microarray based on these sequences was constructed and used in gene expression studies. The differentially expressed genes identified by the CHO cDNA microarray were compared to those observed in cross-species hybridization to mouse cDNA microarrays. Sequence comparison with known mouse genes indicates only a moderate degree of homology. The results represent a significant step toward large-scale gene expression profiling for industrial mammalian cell culture processes.


Biotechnology and Bioengineering | 2008

Genomic and proteomic exploration of CHO and hybridoma cells under sodium butyrate treatment

Joon Chong Yee; Marcela De Leon Gatti; Robin Philp; Miranda Yap; Wei Shou Hu


Biotechnology and Bioengineering | 2005

EST Sequencing for Gene Discovery in Chinese Hamster Ovary Cells

Katie F. Wlaschin; Peter Morin Nissom; Marcela De Leon Gatti; Peh Fern Ong; Sanny Arleen; Kher Shing Tan; Anette Rink; Breana Cham; Kathy Wong; Miranda Yap; Wei Shou Hu


Journal of Bioscience and Bioengineering | 2007

Comparative transcriptional analysis of mouse hybridoma and recombinant Chinese hamster ovary cells undergoing butyrate treatment.

Marcela De Leon Gatti; Katie F. Wlaschin; Peter Morin Nissom; Miranda Yap; Wei Shou Hu

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Wei Shou Hu

University of Minnesota

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Gargi Seth

University of Minnesota

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