Applied Biochemistry and Biotechnology

, Volume 179, Issue 1, pp 168–178 | Cite as

Signal Amplification in Field Effect-Based Sandwich Enzyme-Linked Immunosensing by Tuned Buffer Concentration with Ionic Strength Adjuster

  • Satyendra Kumar
  • Narendra Kumar
  • Siddhartha PandaEmail author


Miniaturization of the sandwich enzyme-based immunosensor has several advantages but could result in lower signal strength due to lower enzyme loading. Hence, technologies for amplification of the signal are needed. Signal amplification in a field effect-based electrochemical immunosensor utilizing chip-based ELISA is presented in this work. First, the molarities of phosphate buffer saline (PBS) and concentrations of KCl as ionic strength adjuster were optimized to maximize the GOx glucose-based enzymatic reactions in a beaker for signal amplification measured by change in the voltage shift with an EIS device (using 20 μl of solution) and validated with a commercial pH meter (using 3 ml of solution). The PBS molarity of 100 μM with 25 mM KCl provided the maximum voltage shift. These optimized buffer conditions were further verified for GOx immobilized on silicon chips, and similar trends with decreased PBS molarity were obtained; however, the voltage shift values obtained on chip reaction were lower as compared to the reactions occurring in the beaker. The decreased voltage shift with immobilized enzyme on chip could be attributed to the increased Km (Michaelis–Menten constant) values in the immobilized GOx. Finally, a more than sixfold signal enhancement (from 8 to 47 mV) for the chip-based sandwich immunoassay was obtained by altering the PBS molarity from 10 to 100 μM with 25 mM KCl.


Enzymatic immunosensors Signal amplification Optimized buffer Sandwich immunoassay 



The authors gratefully acknowledge the financial support of the DST Science and Engineering Research Board, India (Grant No. SB/S3/CE/055/2013).


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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  • Satyendra Kumar
    • 1
    • 3
  • Narendra Kumar
    • 2
    • 3
  • Siddhartha Panda
    • 1
    • 2
    • 3
    Email author
  1. 1.Department of Chemical EngineeringIndian Institute of Technology KanpurKanpurIndia
  2. 2.Materials Science ProgrammeIndian Institute of Technology KanpurKanpurIndia
  3. 3.Samtel Centre for Display TechnologiesIndian Institute of Technology KanpurKanpurIndia

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