A Chemiluminescence Based Optical Biosensor Coupled with Immunomagnetic Separation for the Detection of ββ-Glucuronidase from Escherichia coli βββ-Glucuronidase from βββ-Glucuronidase from

Yıl 2016, Cilt: 14 Sayı: 2, 75 - 84, 01.06.2016

Şeref Tağı Arthur G Rand

Öz

A biosensor was developed using a magnetic focusing optical waveguide glass tubular probe for the detection of glucuronidase GUS from Escherichia coli based on immunomagnetic separation and chemiluminescent measurement. The magnetic focusing resulted in a lower chemiluminescent signal than that obtained without magnetic-focusing, mainly due to the diffusion of the chemiluminescent reaction products into the solution rather than concentrating around the GUS enzyme captured by the antibodies on the paramagnetic beads. A microwell format was also tested for the chemiluminescent assay. A linear relationship was observed between log-GUS concentration and log-chemiluminescent signal for the microwell and magnetic focusing formats with a limit of detection as low as 100 and 1000 pg GUS/mL, respectively. The microwell format had the potential to be improved for higher sensitivity. The procedure was simple and rapid, and the whole assay could be completed in 70 min

Anahtar Kelimeler

Glucuronidase, Chemiluminescence, Biosensor, Immunomagnetic separation, Magnetic focusing

Escherichia coli β-Glukuronidaz Enziminin Saptanmasında İmmunomanyetik Ayırmayla Birleştirilmiş Kemiluminesans Temelli Optik Biyosensör

Öz

Bu araştırmada Escherichia coli glukorunidaz GUS enziminin saptanması amacıyla immunomanyetik ayırma ve kemiluminesans yöntemlerini esas alan biyosensör geliştirilmiştir. Biyosensörde, ucunda odaklayıcı bir mıknatıs bulunan ve tüp biçiminde camdan yapılmış ışık dalga iletim kılavuzu kullanılmıştır. Kemiluminesans tepkime ürünleri paramanyetik kürecikler üzerinde antikorlar tarafından yakalanan GUS enziminin çevresine yoğunlaşmak yerine reaksiyon çözeltisine dağılmıştır. Bu nedenle manyetik odaklama yöntemiyle saptanan kemiluminesans sinyali manyetik odaklama olmadan saptanan sinyale göre daha düşük olmuştur. Ayrıca mikrokuyucuk içerisinde de kemiluminesans yöntemi uygulanmıştır. Logaritmik GUS konsantrasyonu ile logaritmik kemiluminesans ölçüm değerleri arasında doğrusal bir ilişki olduğu görülmüş, mikrokuyucuk ve manyetik odaklama yöntemlerinin duyarlılığı sırasıyla 100 ve 1000 pg GUS/mL olarak saptanmıştır. Mikrokuyucuk yönteminin daha fazla geliştirilme potansiyeli olduğu anlaşılmaktadır. Yöntem basit ve hızlı olup tüm işlemler 70 dakika sürmektedir

Anahtar Kelimeler

Glukuronidaz, Kemiluminesans, Biyosensör, İmmünomanyetik ayırma, Manyetik odaklama

Kaynakça

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