Science: Diffractive optical sensor

chemical reaction with the analyte, it might act as a chemical sensor," he says. .masaka and his group present their work in this is- sue ai Analytica...
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News SCIENCE cific bases, removing the modified base from its sugar and phosphodiester bonds, and cleaving the molecule with piperidine. This process is performed on the original temA new type of chemical sensor allows replate molecule, rather than an enzymatic sults to be displayed without the need for a copy of the template as is done in the more detector or electronics. commonly used Sanger sequencing method. Totaro Imasaka of Kyushu University MS has previously been used to detect frag(Japan) had been interested in diffractive ments from the Sanger method, but, to the optical elements because of their ability to best of Chen's knowledge, this is the first project figures using pulsed laser beams as time MS has been used with Maxam-Gilbert in laser light shows. This gave him an idea sequencing. that went beyond the entertainment value. "If beam diffraction appears as the result of a With Maxam-Gilbert sequencing, the chemical reaction with the analyte, it might DNA is cleaved into two fragments. For the act as a chemical sensor," he says. .masaka mass spectra to make any sense, fragments and his group present their work in this ismust be from the same end of the molesue ai Analytical Chemistry (p 2262). cule. In this case, Chen is sequencing from the 5' end. To collect these fragments, the The device that his group developed 5' end is biotinylated, and the appropriate consists of gelatin with thymolphthalein in fragments are captured by streptavidinalternating strips. This pH indicator is coated magnetic beads. The captured fragtransparent at pH < 9.3 and turns blue at ments are released from the beads by hot pH > 10.5. "We have not tried any other pH ammonia treatment and are analyzed by indicators yet," says Imasaka. "However, all Beam diffraction patterns of (a) the bare MALDI-TOFMS. the pH indicators should work well as long sensor, (b) the sensor dipped in distilled as they have an absorption band at the They demonstrated the method with a water, and (c) the sensor dipped in alkallne wavelength of the laser emission." synthetic 30-mer derived from exon 5 of solution. the human p53 gene and with a 60-mer conWhen light passes through the sensor taining 5'-biotin-(GAA)20. The sequence in acidic solution, there is very little diffracImasaka envisions a detector, such as tion. When the sensor is dipped in distilled this one, being used as a CE detector. .Espe- derived from the mass spectra for the 30water, there is little change in the beam cially in a chip-CE system, a sensitive detec- mer matched the sequence of the original template molecule. The mass resolution pattern. However, when the solution is tor is strongly desired. A laser ffuorometric made alkaline with NaOH, each strip condetector is used in most cases. However, it is (m/Am) was poor—in the range of 30. The reaction products for the 60-mer were diffitaining thymolphthalein turns blue. The expensive, and only fluorescent molecules cult to resolve beyond 40 nucleotides. alternation between the clear gelatin strips are detected. The method using beam difChen suspects that the failure to resolve and the blue gelatin strips sets up a perifraction allows, in theory, more sensitive longer fragments was caused by impurities. odic difference in the light intensity transdetection of light-absorbing analytes." mitted through the device, similar to a Celia Henry 'With the MALDI method almost any impurity, such as salt, can cause the mass transmission grating. The sensor then difspectrometer detection efficiency to be fracts the laser light, producing a series of spots. Imasaka expects that the sensor will Going to the M a x a m quite ,flow and the resolution to be poor " he says We are still lookinj? for an efficient be quantitative for pH change but the There's been talk recently of sequencing way to get pure DNA for the mass spectrogroup has not yet performed any quantitathe human genome within the next two to metric detection " tive experiments four years. That time line is not expected to This method has no advantage over include the estimated 10% of the human Although the Sanger method typically other methods of pH determination in genome represented by "difficult" temdelivers cleaner samples, Maxam-Gilbert terms of the actual values. However, it replates—including DNA with secondary sequencing has certain advantages, says quires no detector or electronics. "The adstructures, a high number of repeats, Chen. "With Maxam-Gilbert sequencing, vantage of the present pH sensor is that the and/or a high G-C component. Winston you don't need to worry about faithful repanalytical result can be directly displayed C. H. Chen and co-workers at Oak Ridge lication," he notes. "If DNA has a secondon a screen without using any electronics," National Laboratory are developing a tech- ary structure or is G-C rich, quite often says Imasaka. "We expect to use this sennique that they hope will help with that [the Sanger method] cannot faithfully sor for direct display of the analytical result "tricky" 10%. They describe their work in replicate the original DNA template. With where no electricity can be used or no elec- this issue of Analytical Chemistry (p 2266)6 the Maxam-Gilbert method, you only cut tricity is available." They are using chemical cleavage (the the DNA; you don't replicate it. You just Maxam-Gilbert reaction) combined with cut this whole region of DNA into differThe concept of diffractive optical senMALDI-TOFMS to sequence DNA Maxam- ent sizes." sors is not restricted to pH measurements. Gilbert sequencing works by modifying "All the colorimetric reactions might be A second advantage of Maxam-Gilbert

Diffractive optical sensor

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used for construction of a diffractive optical sensor—for example, a calcium ion sensor or a protein sensor. There is much basic research in similar fields, for example, optical fiber sensors. All of this research might be useful for the construction of a new type of diffractive optical chemical sensor."

Analytical Chemistry News & Features, July 1, 1999