An Effective Cell Coculture Platform Based on the Electrospun Microtube Array Membrane for Nerve Regeneration

2017 ◽  
Vol 204 (3-4) ◽  
pp. 179-190 ◽  
Author(s):  
V. Chia-Hsuan Tseng ◽  
Chee Ho Chew ◽  
Wan-Ting Huang ◽  
Yang-Kao Wang ◽  
Ko-Shao Chen ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Chain Reaction ◽  
Tissue Culture Polystyrene ◽  
Coculture System ◽  
Submicron Scale ◽  
A Cell ◽  
Polymerase Chain ◽  
Transwell System ◽  
Lower Expression ◽  
Microtube Array

Recently, a novel substrate known as an electrospun polylactic acid (PLLA) microtube array membrane (MTAM) was successfully developed as a cell coculture platform. Structurally, this substrate is made up of one-to-one connected, ultrathin, submicron scale fibers that are arranged in an arrayed formation. Its unique structure confers several key advantages which are beneficial in a cell coculture system. In this study, the interaction between rat fetal neural stem cells (NSC) and astrocytes was examined by comparing the outcome of a typical Transwell-based coculture system and that of an electrospun PLLA MTAM-based coculture system. Compared to tissue culture polystyrene (TCP) and Transwell coculture inserts, a superior cell viability of NSC was observed when cultured in lumens of electrospun PLLA MTAM (with supportive immunostaining images). Reverse transcription polymerase chain reaction revealed a strong interaction between astrocytes and NSC through a higher expression of doublecortin and a lower expression of nestin. These data demonstrate that MTAM is clearly a better coculture platform than the traditional Transwell system.

scholarly journals Altered expression of small leucine-rich proteoglycans (Decorin, Biglycan and Lumican): Plausible diagnostic marker in urothelial carcinoma of bladder

Tumor Biology ◽  
2017 ◽  
Vol 39 (5) ◽  
pp. 101042831769911 ◽  
Author(s):  
Sandeep Appunni ◽  
Vivek Anand ◽  
Madhuram Khandelwal ◽  
Amlesh Seth ◽  
Sandeep Mathur ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Bladder Cancer ◽  
Urothelial Carcinoma ◽  
Messenger Rna ◽  
Quantitative Polymerase Chain Reaction ◽  
Rna Expression ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Lower Expression ◽  
Urothelial Carcinoma Of Bladder

Small leucine-rich proteoglycans are components of extracellular matrix that regulates neoplastic transformation. Among small leucine rich proteoglycans, Decorin, Biglycan and Lumican are most commonly implicated markers, and their expression is well studied in various malignancies. In this novel study, we have collectively evaluated expression of these three molecules in urothelial carcinoma of bladder. Thirty patients of confirmed untreated bladder cancer, 30 healthy controls for blood and 30 controls for adjacent non-tumour tissue were enrolled. Blood was collected from all subjects and tumour/adjacent normal tissue was obtained from the patients. Circulatory levels were estimated by enzyme-linked immunosorbent assay, relative messenger RNA expression by quantitative polymerase chain reaction and protein expression by immunohistochemistry and western-blotting. Circulatory levels of Biglycan (p = 0.0038) and Lumican (p < 0.0001) were significantly elevated, and that of Decorin (p < 0.0001) was significantly reduced in patients as compared with controls. Protein expression by immunohistochemistry and western-blotting showed elevated expression of Lumican and Biglycan and lower expression of Decorin in urothelial carcinoma of bladder. Quantitative polymerase chain reaction for messenger RNA expression from tissue specimens revealed significantly higher expression of Biglycan (p = 0.0008) and Lumican (p = 0.01) and lower expression of Decorin (p < 0.0001) in urothelial carcinoma of bladder. Out of all molecules receiver operating characteristic curve showed that the 0.207 ng/ml cut-off of serum Lumican provided optimum sensitivity (90.0%) and specificity (90.0%). Significant alteration of matrix small leucine-rich proteoglycans in urothelial carcinoma of bladder was observed. Higher expression of Lumican in Bladder cancer patients with the cut-off value of highest optimum sensitivity and specificity shows its importance as a potential non-invasive marker for early detection of UBC following further validation in large patient cohort.

scholarly journals HIV-1 Preintegration Complexes Preferentially Integrate into Longer Target DNA Molecules in Solution as Detected by a Sensitive, Polymerase Chain Reaction-based Integration Assay

2001 ◽  
Vol 276 (50) ◽  
pp. 46946-46952 ◽  
Author(s):  
Alexei Brooun ◽  
Douglas D. Richman ◽  
Richard S. Kornbluth
Keyword(s):  
Polymerase Chain Reaction ◽  
Host Cell ◽  
Polymerase Chain Reaction Amplification ◽  
Chain Reaction ◽  
Preintegration Complex ◽  
Target Dna ◽  
Host Cell Factors ◽  
A Cell ◽  
Polymerase Chain ◽  
Exonuclease Digestion

After entering a cell and undergoing reverse transcription, the retroviral genome is contained in a preintegration complex (PIC) that mediates its integration into host cell DNA. PICs have been shown to prefer torsionally strained DNA, but the effect of target DNA length has not been previously examined. In this report, concatemerization of a repeating 105-base pair unit was used to vary target DNA length independently from basic DNA sequence, while maintaining both PICs and target DNAs in solution. Integration junctions were quantified by real-time fluorescence-monitored polymerase chain reaction amplification using primers in the viral long terminal repeat and the target DNA. Unreacted target DNA severely inhibited the post-reaction polymerase chain reaction detection step, requiring its removal using λ exonuclease digestion. Integration into a 32-unit concatemer of target DNA was markedly more efficient than integration into a monomeric unit, indicating that longer target DNA was preferred. This substrate was used to construct a simple, robust, and adaptable assay that can serve as a method for studying the host cell factors that enhance PIC integration, and as a drug discovery platform for integration inhibitors active against PICs.

Junctions of the AML1/MTG8(ETO) fusion are constant in t(8;21) acute myeloid leukemia detected by reverse transcription polymerase chain reaction

Blood ◽  
1993 ◽  
Vol 82 (4) ◽  
pp. 1270-1276 ◽  
Author(s):  
T Kozu ◽  
H Miyoshi ◽  
K Shimizu ◽  
N Maseki ◽  
Y Kaneko ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Acute Myeloid Leukemia ◽  
Cell Line ◽  
Reverse Transcription ◽  
Myeloid Leukemia ◽  
Chain Reaction ◽  
Translocation Breakpoints ◽  
A Cell ◽  
Polymerase Chain ◽  
Acute Myeloid

Abstract The chromosomal translocation, t(8;21), is found frequently in acute myeloid leukemia (AML) with maturation (FAB-M2). We have previously mapped the translocation breakpoints of t(8;21) in a specific intron of the AML1 gene on chromosome 21. In this study, we cloned cDNAs synthesized from a cell line carrying t(8;21) by reverse transcription polymerase chain reaction (RT-PCR) using an AML1-specific primer. The analysis of the cDNAs structure has led to the identification of the fusion of AML1 with a gene named MTG8 on chromosome 8, which seems to be identical to ETO. Northern analysis using MTG8 (ETO) probes detected 7.8-kb and 6.2-kb RNAs and several minor RNAs in the cell line with t(8;21), but failed to detect any transcripts in a cell line without t(8;21). A set of primers were designed to detect the AML1/MTG8(ETO) fusion by PCR. The PCR amplified identical products in all 6 patients and one cell line with t(8;21), suggesting that the AML1/MTG8(ETO) fusion is a constant feature associated with t(8;21) and the junctions of the AML1/MTG8(ETO) fusion are restricted in a unique site. Because the PCR detection of the AML1/MTG8(ETO) fusion at the RNA level is highly sensitive, it can be used as a sensitive method for diagnosis and detection of minimal residual disease in t(8;21) leukemia.

scholarly journals Detection of ureaplasma urealyticum by polymerase chain reaction examination in nonspecific genital infection patients

2019 ◽  
Author(s):  
R. Rahmadewi ◽  
Dian PH
Keyword(s):  
Polymerase Chain Reaction ◽  
Ureaplasma Urealyticum ◽  
Genital Infection ◽  
Chain Reaction ◽  
Sexually Transmitted ◽  
Drug Of Choice ◽  
High Incidence ◽  
A Cell ◽  
Pcr Method ◽  
Polymerase Chain

Non specific genital infection (NSGI) is a condition affecting females which causes inflammation of the endocervix or anterior urethra that is not caused by Neisseria gonorrhoeae. The causative sexually transmitted organisms include Chlamydia trachomatis (Groups D to K) and Ureaplasma urealyticum. Infection caused by Ureaplasma urealyticum is often asymptomatic even though many studies have pronounced that Ureaplasma urealyticum can contribute not only to lower genitourinary infection but also to infertility. Ureaplasma urealyticum cannot be stained by Gram stain due to the lack of a cell wall of the organism. This research aims to evaluate the prevalence of Ureaplasma urealyticum in NSGI patients by using the polymerase chain reaction (PCR) method targeted in the ureaplasma gene structure 429 bp area. The samples were extracted from eighteen DNA NSGI patients. Eleven out of eighteen (61.11%) DNA NSGI samples tested positive for Ureaplasma urealyticum. Most patients (44.44%) with Ureaplasma urealyticum were unemployed, and 27.78% were complaining of recurrent vaginal discharge. The high incidence of Ureaplasma urealyticum in this study needs further attention since doxycycline remains the drug of choice of NSGI. Moxifloxacin should be considered for patients who are making no clinical progress with doxycycline.

scholarly journals Junctions of the AML1/MTG8(ETO) fusion are constant in t(8;21) acute myeloid leukemia detected by reverse transcription polymerase chain reaction

Blood ◽  
1993 ◽  
Vol 82 (4) ◽  
pp. 1270-1276 ◽  
Author(s):  
T Kozu ◽  
H Miyoshi ◽  
K Shimizu ◽  
N Maseki ◽  
Y Kaneko ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Acute Myeloid Leukemia ◽  
Cell Line ◽  
Reverse Transcription ◽  
Myeloid Leukemia ◽  
Chain Reaction ◽  
Translocation Breakpoints ◽  
A Cell ◽  
Polymerase Chain ◽  
Acute Myeloid

The chromosomal translocation, t(8;21), is found frequently in acute myeloid leukemia (AML) with maturation (FAB-M2). We have previously mapped the translocation breakpoints of t(8;21) in a specific intron of the AML1 gene on chromosome 21. In this study, we cloned cDNAs synthesized from a cell line carrying t(8;21) by reverse transcription polymerase chain reaction (RT-PCR) using an AML1-specific primer. The analysis of the cDNAs structure has led to the identification of the fusion of AML1 with a gene named MTG8 on chromosome 8, which seems to be identical to ETO. Northern analysis using MTG8 (ETO) probes detected 7.8-kb and 6.2-kb RNAs and several minor RNAs in the cell line with t(8;21), but failed to detect any transcripts in a cell line without t(8;21). A set of primers were designed to detect the AML1/MTG8(ETO) fusion by PCR. The PCR amplified identical products in all 6 patients and one cell line with t(8;21), suggesting that the AML1/MTG8(ETO) fusion is a constant feature associated with t(8;21) and the junctions of the AML1/MTG8(ETO) fusion are restricted in a unique site. Because the PCR detection of the AML1/MTG8(ETO) fusion at the RNA level is highly sensitive, it can be used as a sensitive method for diagnosis and detection of minimal residual disease in t(8;21) leukemia.

Detection of poliovirus by cell culture and by PCR after UV disinfection

1995 ◽  
Vol 31 (5-6) ◽  
pp. 141-145 ◽  
Author(s):  
A. Maier ◽  
D. Tougianidou ◽  
A. Wiedenmann ◽  
K. Botzenhart
Keyword(s):  
Cell Culture ◽  
Polymerase Chain Reaction ◽  
Uv Irradiation ◽  
Viral Rna ◽  
Uv Disinfection ◽  
Mercury Lamp ◽  
Chain Reaction ◽  
A Cell ◽  
Polymerase Chain ◽  
Cell Culture Assay

The polymerase chain reaction (PCR) is used for the detection of microorganisms in water, but it is known that one does not detect only viable microorganisms by PCR. The following results show the detection of poliovirus on a cell culture assay and by PCR after inactivation by UV irradiation. The virus was irradiated with a low pressure mercury lamp with times up to 4000 seconds (=3980 J/m2). A 4 log inactivation of the virus was complete after 300 seconds (=294 J/m2) as shown by the cell culture assay. The viral RNA was detectable by PCR even at irradiation times of 4000 seconds.

Detection by Polymerase Chain Reaction of all Common Mycoplasma in a Cell Culture Facility

Pathobiology ◽  
1995 ◽  
Vol 63 (1) ◽  
pp. 9-11 ◽  
Author(s):  
James M. Pruckler ◽  
Janet M. Pruckler ◽  
Edwin W. Ades
Keyword(s):  
Cell Culture ◽  
Polymerase Chain Reaction ◽  
Chain Reaction ◽  
A Cell ◽  
Polymerase Chain ◽  
Cell Culture Facility

High performance Nanogold™-in situ hybridzation and its use in the detection of hybridized and PCR-amplified microscopical preparations

1996 ◽  
Vol 54 ◽  
pp. 896-897
Author(s):  
G. W. Hacker ◽  
I. Zehbe ◽  
J. Hainfeld ◽  
A.-H. Graf ◽  
C. Hauser-Kronberger ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Messenger Rna ◽  
High Performance ◽  
Detection System ◽  
Direct Methods ◽  
Genetic Alterations ◽  
Chain Reaction ◽  
Protein Encoding ◽  
Polymerase Chain

In situ hybridization (ISH) with biotin-labeled probes is increasingly used in histology, histopathology and molecular biology, to detect genetic nucleic acid sequences of interest, such as viruses, genetic alterations and peptide-/protein-encoding messenger RNA (mRNA). In situ polymerase chain reaction (PCR) (PCR in situ hybridization = PISH) and the new in situ self-sustained sequence replication-based amplification (3SR) method even allow the detection of single copies of DNA or RNA in cytological and histological material. However, there is a number of considerable problems with the in situ PCR methods available today: False positives due to mis-priming of DNA breakdown products contained in several types of cells causing non-specific incorporation of label in direct methods, and re-diffusion artefacts of amplicons into previously negative cells have been observed. To avoid these problems, super-sensitive ISH procedures can be used, and it is well known that the sensitivity and outcome of these methods partially depend on the detection system used.

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