Highly alkaline electrolyte for single-stranded DNA separations by electrophoresis in bare silica capillaries

Malá, Zdeňka; Klepárnı́k, Karel; Boček, Petr

doi:10.1016/s0021-9673(99)00476-8

Cited by 27 publications

(16 citation statements)

References 36 publications

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“…This was despite a lower absolute AA concentration in the PEG:AA:DiPEG grafting mixture since the total monomer concentration was maintained constant rather than the concentration of any one mono- mer (Table 1). To explain this result, recall that m eo is given by ez/Z where e is the dielectric constant, z is the zeta potential of the liquid-solid interface, and Z is the viscosity of the solution near the solid surface [32]. It is likely that the cross-linked PEG:AA forms a nearly solid surface.…”

Section: Characterization Of Pdms Surfaces Grafted With Peg Aa and mentioning

confidence: 99%

Cross‐linked coatings for electrophoretic separations in poly(dimethylsiloxane) microchannels

Hu¹,

Ren²,

Bachman

et al. 2003

Electrophoresis

100

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We have developed a strategy using ultraviolet light to polymerize mixed monomer solutions onto the surface of a poly(dimethylsiloxane) (PDMS) microdevice. By including monomers with different chemical properties, electrophoretic separations were optimized for a test set of analytes. The properties of surfaces grafted with a single neutral monomer, a neutral and a negative monomer, or a neutral, negative, and cross-linking monomer were assessed. The highest quality separations were achieved in channels with cross-linked coatings. The separation efficiency for biologically relevant peptides (kinase substrates) on these surfaces was as high as 18 600 theoretical plates in a 2.5 cm channel. The test peptides were fluorescein-AEEEIYGEFEAKKKK, fluorescein-GRPRAATFAEG, fluorescein-GRPRAA(T-PO(3))FAEG, fluorescein-DLDVPIP GRFDRRVSVAAE, and fluorescein-DLDVPIPGRFDRRV(S-PO(3))VAAE. Separations between two different peptides occurred in as little as 400 ms after injection into the separation channel. The simultaneous separation of five kinase and phosphatase substrates was also demonstrated. By carefully selecting mixtures of monomers with the appropriate properties, it may be possible to tailor the surface of PDMS for a large number of different electrophoretic separations.

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Section: Characterization Of Pdms Surfaces Grafted With Peg Aa and mentioning

confidence: 99%

Cross‐linked coatings for electrophoretic separations in poly(dimethylsiloxane) microchannels

Hu¹,

Ren²,

Bachman

et al. 2003

Electrophoresis

100

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“…In general, a highly alkaline environment can convert double-stranded chromosomal DNA into ssDNA (Mala et al 1999). Moreover, it has been reported that a surprisingly large number of ssDNA breaks can be detected after high-alkaline treatment (Singh et al 1989); thus, the assessment of the chromosomal DNA integrity of spermatozoa treated with various concentrations of NaOH becomes crucial.…”

Section: Chromosome Damage In Naoh-treated Spermatozoamentioning

confidence: 99%

Production of normal mice from spermatozoa denatured with high alkali treatment before ICSI

Li¹,

Mizutani²,

Ono³

et al. 2009

Reproduction

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In mammals, ICSI is now a very important tool for both assisted reproductive technology and studying the mechanisms of fertilization. In the latter experiments, it is important to use spermatozoa that have lost their oocyte activation capacity but still retain their developmental potential. In this study, we used high-concentration NaOH to remove oocyte activation potential from spermatozoa, and examined whether normal offspring could be generated from these spermatozoa after ICSI. The spermatozoa were treated with different concentrations of NaOH (1-100 mM) for 1 h and then neutralized with equal amounts of same concentration of HCl. In 10 mM NaOHtreated spermatozoa, the cell membrane was broken and most of them failed to activate oocytes after their injection into the oocytes. However, these spermatozoa did not show strong damage, and after artificial activation with SrCl 2 , all of the zygotes were judged as normal by immunostaining to check the methylation status of histone H3 lysine 9, low chromosome damage by karyotype assay and staining with DNA double-strand breaks marker, gH2AX. Moreover, after transferring those embryos into recipient females, 106 (36.7%) live and healthy offspring were delivered, which is similar to the rate in the fresh control group. By contrast, spermatozoa treated with lower NaOH concentrations retained their oocyte activation capacity and those treated with higher concentrations lost their developmental potential. This suggests that 10 mM NaOH for 1 h is the best treatment to completely destroy the cell membrane and activation capacity of spermatozoa without injuring their developmental potential.

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“…4 and 5), which is particularly sensitive to temperature variation to demonstrate that their CE systems are stable and well calibrated [47]. CE analysis of DNA fragments at elevated pH conditions, where the DNA molecule is predominately denatured, suggests that DNA secondary structure is responsible for the variations observed in DNA size determinations with fluctuating temperatures [48][49][50]. By carefully controlling the run conditions, i.e., pH, buffer, denaturants, and temperature, variations within and between runs can be minimized and overall run precision improved.…”

Section: The Buffermentioning

confidence: 99%

Forensic DNA typing by capillary electrophoresis using the ABI Prism 310 and 3100 genetic analyzers for STR analysis

et al. 2004

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DNA typing with short tandem repeat (STR) markers is now widely used for a variety of applications including human identification. Capillary electrophoresis (CE) instruments, such as the ABI Prism 310 and ABI 3100 Genetic Analyzers, are the method of choice for many laboratories performing STR analysis. This review discusses issues surrounding sample preparation, injection, separation, detection, and interpretation of STR results using CE systems. Requirements for accurate typing of STR alleles are considered in the context of what future analysis platforms will need to increase sample throughput and ease of use.

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Highly alkaline electrolyte for single-stranded DNA separations by electrophoresis in bare silica capillaries

Cited by 27 publications

References 36 publications

Cross‐linked coatings for electrophoretic separations in poly(dimethylsiloxane) microchannels

Cross‐linked coatings for electrophoretic separations in poly(dimethylsiloxane) microchannels

Production of normal mice from spermatozoa denatured with high alkali treatment before ICSI

Forensic DNA typing by capillary electrophoresis using the ABI Prism 310 and 3100 genetic analyzers for STR analysis

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