Microchip electrophoresis is a technique used to separate proteins, nucleic acids, and charged molecules in an electric field, which enhances speed and simplicity compared to traditional methods. While it allows rapid separation and automation, it faces challenges such as limited separation efficiency and low sensitivity in detection. The technology involves the creation of microchips with precise channel designs and detection methods like laser-induced fluorescence, aiming for cost-effective and efficient biochemical analysis.

1. MICROCHIP
ELECTROPHORESIS
- Dr.Arun Babu.N.B
II year MD (Biochemistry)
V.M.K.V.Medical College

2. INTRODUCTION
DEFINITION
Technique used to separate various proteins, nucleic
acids and other charged molecules under the influence of
an electric field.
PRINCIPLE
In an electric field, the charged molecules move towards
oppositely charged electrode at different rates based on
electrical charge & molecular size.

3. 1st
Electrophoresis method- to study proteins- was Free
solution/ moving boundary method- By TISELIUS
It was used to measure electrophoretic mobility and to study
protein-protein interaction.
It was able to resolve serum proteins into 4 component
mixtures, with α1 fraction incompletely separated from
albumin.

4. An ampholyte/ zwitterion becomes +vely charged in a
solution that is more acidic than its isoelectric point, and
migrates to cathode and vice versa.
Electrophoretic mobility is directly proportional to net
charge & inversely proportional to molecular size and
viscosity of the electrophoresis medium.

5. Mobility may be +ve or -ve, depending on whether the
protein migrates in the same or opposite direction as the
electrophoretic field, which is from anode to cathode.
Factors affecting electrophoresis are:
Electric Field (voltage, current, resistance)
Sample (charge, size, shape)
Buffer (composition, concentration, pH)
Supporting medium (electroendosmosis)

6. TYPES
Based on nature of supporting medium
AGE (Agarose gel electrophoresis)
PAGE (Poly-acrylamide gel electrophoresis)
Paper strip electrophoresis (cellulose acetate paper/membrane)
Based on mode of technique
Slide/Slab gel electrophoresis
Disc electrophoresis
Isoelectric focusing electrophoresis
Capillary electrophoresis
Microchip electrophoresis …..

7. MICROCHIP
ELECTROPHORESIS
Recently undergone substantial development like
integrated microchip designs, advanced direction systems
DNA & proteins
ADVANTAGES
High Speed
4x – 10x faster than conventional capillary electrophoresis
1 order of magnitude faster than slab gel electrophoresis
Simplicity
Potential for automation

8. DISADVANTAGES
Limited separation efficiency of zone electrophoretic
measurements
Imprecise injection
Low sensitivity of absorption detection(UV/Vis
absorption detection)
Early stage of commercialization

9. INSTRUMENTATION
Separation channels
Sample injection channels
Reservoirs
Sample preparation reactors
Pre and post column reactors
Truly multi-functional, “integrated” analytical device
embedded in a single monolithic substrate.
Fabricated onto
the surface of the
microchip, using
photolithographic
processes.

10. Usually cross T design [double T(larger injector region)]
Has a short (injection) channel & longer (separation)
channel
1 reservoir each at each end
2 for introduction of sample & background electrolyte (buffer
solution)
2 serving as waste reservoirs
Channel dimensions (depth=15-50μm, width=50-200μm
and length of separation channel=1-10cm)

11. The volume of the separation channel is 1 order of
magnitude smaller than conventional capillary systems.
Sample volume injected varies from 100-500pL.
With the decrease in volume requirements, pressure
injection is more challenging. Hence sample is injected
electro kinetically, by applying an electric field across the
sample channel.

12. All reservoirs are connected to electrodes
An injection voltage of several 100V is applied across the
sample and sample waste reservoirs- to migrate the
sample to the injection cross
Separation voltage (1-4kV) is then applied to separation
channel, which induces the separation of analyte zones
before they reach detection window downstream.

13. Portion of the sample present in the intersection
represents the injection plug, which is subjected to
separation when electric field is applied across separation
channel
Detection on microchip is usually made at opposite end
of separation channel, most commonly by LIF (Laser
Induced Fluorescence) due to its sensitivity.
Typical microchip separation time- 50-200seconds.

17. DETECTION
LIF (Laser Induced Fluorescence) – Most commonly
used method on chip due to its high sensitivity.
Most analytes are not fluorophores & have to be derivatized to
be detected by LIF.
LIF is much larger than the microfabricated separation device,
which makes it unfavorable for portable analytical device.
Electrochemical detection
Amperometric detection
Voltametric detection
Conductiometric detection
Potentiometric detection

19. APPLICATIONS
For simultaneous separation of catecholamines & their
cationic metabolites.
To enhance sensitivity of on-chip amperometric detection
Carbon nanotube modified amperometry
Microchip Affinity Capillary electrophoresis (MC-ACE)
For Enzyme assays
Microchip isoelectric Focusing (MC-IEF)
To compare practical applicability of pharmaceuticals

20. FABRICATION
Microchips are constructed from substrates such as:
Glass (Pyrex-like or soda lime)
Silicon (as per microelectronic chips)
Polymeric materials (plastics)
Silicon-like materials (polydimethylsiloxane)

22. A buffered solution of HCl is used to etch the desired
structures into a glass wafer, thereby producing a series
of U-shaped troughs that interconnect appropriately.
Channels are U-shaped because of downward & lateral
etching by the etch solution.
After etching, the etched wafer is bonded to a 2nd
piece of
glass, into which reservoirs have been drilled, to enclose
the chambers and channels of the device.

25. REFERENCES
Tietz Textbook of Clinical Chemistry & Molecular Diagnostics; 5th
Edition
Microchip Capillary Electrophoresis-Methods & protocols:
Charles.S.Henry; 2006 edn
S. Gotz, U. Karst, Anal. Bioanal. Chem. 387 (2007) 183
W.R.Vandaveer, S.A.Pasas-Farmer, D.J.Fischer, C.N.Frankenfeld,
S.M.Lunte, Electrophoresis 25 (2004) 3528
M.A. Schwarz, P.C. Hauser, Lab Chip 1 (2001) 1
E.S. Roddy, H.W. Xu, A.G. Ewing, Electrophoresis 25 (2004) 229
www.micruxfluidic.com/archivos/videos/micrux_mce.swf

26. Thank
You