Microplate-based fluorometric methods for the enzymatic determination of l-glutamate: application in measuring l-glutamate in food samples

Chapman, Justin; Zhou, Mingjie

doi:10.1016/s0003-2670(99)00533-4

Cited by 61 publications

(32 citation statements)

References 11 publications

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“…3a), although under these conditions, xanthine concentrations >1.2 M would need a different instrumental setup and higher amounts of Amplex Red. The lower detection limit, calculated following IUPAC recommendations (Thomsem et al, 2003), was also the same for both systems (20 nM), and similar to that determined for other Amplex Red-based enzymatic assays (Chapman and Zhou, 1999;MartinezPérez et al, 2003;Pastor et al, 2004). This value largely improves the sensitivity of the currently xanthine biosensors, as is shown in Table 1, and allows to handle diluted samples to further reduce the presence of interference substances (Martinez-Pérez et al, 2003).…”

Section: Enzyme Activity and Characterization Of Sensor Performancesupporting

confidence: 76%

Immobilization of a trienzymatic system in a sol–gel matrix: A new fluorescent biosensor for xanthine

Salinas-Castillo

Pastor

Mallavia

et al. 2008

Biosensors and Bioelectronics

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Section: Enzyme Activity and Characterization Of Sensor Performancesupporting

confidence: 76%

Immobilization of a trienzymatic system in a sol–gel matrix: A new fluorescent biosensor for xanthine

Salinas-Castillo

Pastor

Mallavia

et al. 2008

Biosensors and Bioelectronics

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“…The activity of GLNase could not be analyzed with the existing cycling systems. Several methods to determine the reaction product Glu are based on the enzymatic recycling of the substrate (Glu), for example spectrophotometric methods (Valero and Garcia-Carmona, 1998) or fluorometric methods (Chapman and Zhou, 1999). To avoid fluorometric analysis and to use a similar approach as for the other 27 enzymes described here, a new GLNase assay is proposed which is based on substrate cycling between GLDH and AspTA.…”

Section: Resultsmentioning

confidence: 99%

Measurement of key metabolic enzyme activities in mammalian cells using rapid and sensitive microplate‐based assays

Janke

Genzel

Wahl

et al. 2010

Biotech & Bioengineering

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Sensitive microplate-based assays to determine low levels of key enzyme activities in mammalian cells are presented. The enzyme platform consists of four cycling assays to measure the activity of 28 enzymes involved in central carbon and glutamine metabolism. The sensitivity limit of all cycling assays was between 0.025 and 0.4 nmol product. For the detection of glutaminase activity, a new glutamate cycle system involving the enzymes glutamate dehydrogenase and aspartate transaminase was established. The relative standard deviation of the method was found to be 1.7% with a limit of detection of 8.2 pmol and a limit of quantitation of 24.8 pmol. Hence, cell extracts could be highly diluted to reduce interferences caused by other components in the extract, which in addition minimized underestimates or overestimates of actual enzyme activities. Since substrate concentrations could be maintained at a nearly constant level throughout the assay product accumulation during the reaction was low, which minimized product inhibition. As an example, the enzyme platform was used to investigate maximum enzyme activities of stationary-phase MDCK cells grown in serum-containing GMEM medium as typically used in influenza vaccine production.

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“…The latter, based on a fluorimetric microtiter plate assay described by Chapman & Zhou [11] which has become the basis for commercial assays with Amplex Red and HRP (by Molecular Probes, Eugene, OR), was previously validated for use with EBC samples by our group (data not shown). The correlation of data points was easy, as the fluorimetric and enzymatic/amperometric flow injection method differed only in the readout method, i.e., the same samples were analyzed with both methods.…”

Section: Detection Of H 2 O 2 In Exhale Breath Condensatesmentioning

confidence: 99%

Novel Electrochemical Assay for H2O2 Determination in Aqueous Solutions: A Non Time-Critical Method for H2O2 Trace Level Detection

Gajovic‐Eichelmann

Bier

2005

Electroanalysis

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A two-step method for H 2 O 2 trace level determination has been developed, utilizing, for the first time, the fluorogenic peroxidase substrate, Amplex Red (N-acetyl-3,7-dihydroxyphenoxazine), in conjunction with an electroanalytical detection scheme: Upon peroxidase catalyzed turnover of H 2 O 2 with Amplex Red, the fluorescent and electroactive product resorufin (7-hydroxy-3H-phenoxazin-3-one) was detected amperometrically in a thin-layer flow cell with high selectivity. A linear detection range from 100 nmol L À1 through 1000 nmol L À1 was obtained. The performance was validated using exhaled breath condensates (EBC) with and without H 2 O 2 added, as real samples. Due to the improved chemical stability of resorufin as compared to H 2 O 2 , processing larger numbers of EBC samples is greatly facilitated with the new assay, since the enzymatic reaction can be performed up to 18 hours before the actual measurement.

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Microplate-based fluorometric methods for the enzymatic determination of l-glutamate: application in measuring l-glutamate in food samples

Cited by 61 publications

References 11 publications

Immobilization of a trienzymatic system in a sol–gel matrix: A new fluorescent biosensor for xanthine

Immobilization of a trienzymatic system in a sol–gel matrix: A new fluorescent biosensor for xanthine

Measurement of key metabolic enzyme activities in mammalian cells using rapid and sensitive microplate‐based assays

Novel Electrochemical Assay for H2O2 Determination in Aqueous Solutions: A Non Time-Critical Method for H2O2 Trace Level Detection

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