Biochemical SocietyTransactions( 1 992) 20 1 19s High-dielectric media for cell electromanipulation and for electro-kinetic methods
W. Michael ARNOLD Dept. of Biotechnology, University of Wurzburg Rhtgenring 11, D-W8700 Wiirzburg, Germany Methods that use electrical fields to move, separate, or measure cells or macromolecules are on the increase. Cell positioning by dielectrophoresis [l] is now much used in electrofusion [2]. Dielectrophoresis [ 3 ] and electro-rotation [ 4 , 5 3 are established as cell-measurement methods. The use of highfrequency travelling-waves produced by microfabricated structures has been proposed for the handling of cells [61, so that fully electrical cell-sorters can now be envisaged. Many of these methods suffer from the heating and membrane breakdown that result from the application of high fields. These may be avoided if the permittivity of the medium can be increased as described here. TABLE 1 Some electric-field-driven Drocesses Essent i a1 Effect Frequency range Min. Max. Dararneter ( 5 ) Electrophoresis OHz clkHz g/9 Dielectrophoresis lkHz (lGHz 8/9, (t~-Ca)/r) Cell-cell force & Levitation lkHz clGHz C, 8P-CM Elect rolOOHz (lGHz e/9, (CP-~M)/~ rotation Travellingwave drive lOOHz < lGHz C / 9 , (eP-tn)/V In all these methods, the permittivity 8 (so-called dielectric constant or DK) of the medium is very important. The electrophoretic force is directly proportional to the DK, whilst in methods where high frequencies are used (so high that conductive effects are small), the difference between the DK's of particle and medium ( t p - 8 ~ in Table 1) has an additional multiDlicative effect. In these cases, increase of the medium DK can produce an over-linear increase in the electrical force, or even reverse it. A further important parameter considered in Table 1 is the medium viscosity ( 9 ) . Techniques which involve particle motion (not just the exertion of a force) require the lowest possible 9 . It is little appreciated that a variety of materials can increase the (already high) DK of water when dissolved or dispersed in it. A selection of these are shown in Table 2: only those substances with reasonable physiological compatibility are included. The parameters Y and 6 are the DK increments per gram/l and per mole/l solution respectively. Due to the time required for the polarization effects involved, there exists for each substance some frequency f o where the DK increment has fallen to half its low frequency value. (This limits the use of that substance: in addition the conductivity increases in such a "dispersion" region). The last parameter in Table 2 assesses how good a given solute will be at increasing the speed of particles subject to a given field strength (at frequencies well below fo): 8 = ( D K L /DKo - 1 ) / (nL/no - 1 ) where subscripts L and 0 refer to solution and water respectively.
TABLE 2 Parameters of hish-dielectric substances (amino acids. proteins. colloids) in water ViscosityPermittivity increment Critical corrected per: g/l mole/l frequency potency 7
d
amino acids [7] glycine 0.30 22.6 B-alanine 34.6 -NHn caproic acid 77.5 peptides [7,8] gly-gly 0.53 70.6 glY-glY-glY 0.60 113 lys-glu 1.25 345 Protein [9] B- Lact o1.51 60500 globulin Latex particles [ l o ] . 0 8 8 ~ mdiameter 1.8 -.566pm diameter 0.2 -1 . 1 7 ~ ~diameter 1 33 -Polyglutamic acid PGA, MW 90k [ll] 460 -DNA, (calf thymus) lmR NaCl [12] 4500 -Low-salt [12] 80000 -Viscosities: "C" from Ref. (S+Ml; "*" fro@ the Einstein relation
fo
e
3.2GHz 2.4GHz
1.86f 1.71C
l.2GHz
1.80L
l.4GHz 880MHz 370MHz
2.44
2.4MHz
3.14$
3.61L
80kHz 15kHz O.5kHz
9" 41* 165"
16kHz
?
20Hz
?
2Hz ? "$" frm Ref.h3]
Only those materials with fo values of lOOMHz or higher can be used over the full range of the methods shown in Table 1. This restricts the choice to amino acids and short peptides, which at concentrations of 1-3 mole/l are capable of giving DK values of 300 or more (about 4 times that of water). In terms of both 8 and 8 , the best known material is lysylglutamate. However, this tetrapole [ 8 ] is expensive; materials such as glycylglycine are more practical. Amino acids and peptides owe their properties to their zwitterionic nature, the low net charge of which contributes little to the conductivity. This is important in reducing the heat produced in the presence of typical electric fields of 100-500 V/cm. Proteins are poorer in this respect, and most have low dielectric increments per gram. The remaining substances in Table 2 are polyanions, and will give rise to substantial conductivities. Their polarization mechanisms are slow or very slow. Nevertheless, their very high increments per gram, as well as the large values for 8 of the latex particles make them very attractive for low-frequency work (e.g. electrophoresis, provided that their own mobility is not a problem). 1. Pohl,H.A. (1979) Melectrophoresis, University Press, -ridge 2. Z ~ m m , l J . (1982) Biochjm. Riophys. Acta 694, 227-277 3. Kaler,K.V.I.S and Jaw?s,T.B. (1990) Biophys. J. 57, 173-182 4. Amold,W.M. and 2 i m m . U . (1988) J. Electwtat. P, 151-191 5. Fuhr,G., Glaser,R.and Hagedorn,R. (1986) Bi0phys.J. 49,395402 6. Fuhr,G., Hagedorn,R., MUller,T., Wagner,B. and Benecke,W. (1991) Proc. NicmElectro-Memanica~ System Ccng., Nara. 259-264 7. Wyman,J. and Mdh?kin,T.L.(1933)J. Am. Qlem. Soc. 55, 908-914 8. Greenstein,J.P., Wyman,J., and Cdm,E.J. (1935) t l . Rol. olern. Soc. 57, 637-642 9 . Ferry,J.D. and Wley,J.L. (1941) J. Am. Qlem. Soc. 63, 272-278 10. Srhwan,H.P., Srhwarz,G., Manuk,J. and Pauly,H. (1962) J. F@s. Qlern. 66,2626-2635 11. Muller,G. Van der Tauw,F. Zwolle,S. and Mande1,M. (1974) Biophys. chem. 2, 242-254 12. Hayakawa,R., Kanda,H., Sakrmot0.M and Wada,Y. (1975) Japnes Journal of Applied physics 14, 2039-2052 13. mlson,~,(1939) blloid Zeitschrift 68, 51
W. Michael ARNOLD Dept. of Biotechnology, University of Wurzburg Rhtgenring 11, D-W8700 Wiirzburg, Germany Methods that use electrical fields to move, separate, or measure cells or macromolecules are on the increase. Cell positioning by dielectrophoresis [l] is now much used in electrofusion [2]. Dielectrophoresis [ 3 ] and electro-rotation [ 4 , 5 3 are established as cell-measurement methods. The use of highfrequency travelling-waves produced by microfabricated structures has been proposed for the handling of cells [61, so that fully electrical cell-sorters can now be envisaged. Many of these methods suffer from the heating and membrane breakdown that result from the application of high fields. These may be avoided if the permittivity of the medium can be increased as described here. TABLE 1 Some electric-field-driven Drocesses Essent i a1 Effect Frequency range Min. Max. Dararneter ( 5 ) Electrophoresis OHz clkHz g/9 Dielectrophoresis lkHz (lGHz 8/9, (t~-Ca)/r) Cell-cell force & Levitation lkHz clGHz C, 8P-CM Elect rolOOHz (lGHz e/9, (CP-~M)/~ rotation Travellingwave drive lOOHz < lGHz C / 9 , (eP-tn)/V In all these methods, the permittivity 8 (so-called dielectric constant or DK) of the medium is very important. The electrophoretic force is directly proportional to the DK, whilst in methods where high frequencies are used (so high that conductive effects are small), the difference between the DK's of particle and medium ( t p - 8 ~ in Table 1) has an additional multiDlicative effect. In these cases, increase of the medium DK can produce an over-linear increase in the electrical force, or even reverse it. A further important parameter considered in Table 1 is the medium viscosity ( 9 ) . Techniques which involve particle motion (not just the exertion of a force) require the lowest possible 9 . It is little appreciated that a variety of materials can increase the (already high) DK of water when dissolved or dispersed in it. A selection of these are shown in Table 2: only those substances with reasonable physiological compatibility are included. The parameters Y and 6 are the DK increments per gram/l and per mole/l solution respectively. Due to the time required for the polarization effects involved, there exists for each substance some frequency f o where the DK increment has fallen to half its low frequency value. (This limits the use of that substance: in addition the conductivity increases in such a "dispersion" region). The last parameter in Table 2 assesses how good a given solute will be at increasing the speed of particles subject to a given field strength (at frequencies well below fo): 8 = ( D K L /DKo - 1 ) / (nL/no - 1 ) where subscripts L and 0 refer to solution and water respectively.
TABLE 2 Parameters of hish-dielectric substances (amino acids. proteins. colloids) in water ViscosityPermittivity increment Critical corrected per: g/l mole/l frequency potency 7
d
amino acids [7] glycine 0.30 22.6 B-alanine 34.6 -NHn caproic acid 77.5 peptides [7,8] gly-gly 0.53 70.6 glY-glY-glY 0.60 113 lys-glu 1.25 345 Protein [9] B- Lact o1.51 60500 globulin Latex particles [ l o ] . 0 8 8 ~ mdiameter 1.8 -.566pm diameter 0.2 -1 . 1 7 ~ ~diameter 1 33 -Polyglutamic acid PGA, MW 90k [ll] 460 -DNA, (calf thymus) lmR NaCl [12] 4500 -Low-salt [12] 80000 -Viscosities: "C" from Ref. (S+Ml; "*" fro@ the Einstein relation
fo
e
3.2GHz 2.4GHz
1.86f 1.71C
l.2GHz
1.80L
l.4GHz 880MHz 370MHz
2.44
2.4MHz
3.14$
3.61L
80kHz 15kHz O.5kHz
9" 41* 165"
16kHz
?
20Hz
?
2Hz ? "$" frm Ref.h3]
Only those materials with fo values of lOOMHz or higher can be used over the full range of the methods shown in Table 1. This restricts the choice to amino acids and short peptides, which at concentrations of 1-3 mole/l are capable of giving DK values of 300 or more (about 4 times that of water). In terms of both 8 and 8 , the best known material is lysylglutamate. However, this tetrapole [ 8 ] is expensive; materials such as glycylglycine are more practical. Amino acids and peptides owe their properties to their zwitterionic nature, the low net charge of which contributes little to the conductivity. This is important in reducing the heat produced in the presence of typical electric fields of 100-500 V/cm. Proteins are poorer in this respect, and most have low dielectric increments per gram. The remaining substances in Table 2 are polyanions, and will give rise to substantial conductivities. Their polarization mechanisms are slow or very slow. Nevertheless, their very high increments per gram, as well as the large values for 8 of the latex particles make them very attractive for low-frequency work (e.g. electrophoresis, provided that their own mobility is not a problem). 1. Pohl,H.A. (1979) Melectrophoresis, University Press, -ridge 2. Z ~ m m , l J . (1982) Biochjm. Riophys. Acta 694, 227-277 3. Kaler,K.V.I.S and Jaw?s,T.B. (1990) Biophys. J. 57, 173-182 4. Amold,W.M. and 2 i m m . U . (1988) J. Electwtat. P, 151-191 5. Fuhr,G., Glaser,R.and Hagedorn,R. (1986) Bi0phys.J. 49,395402 6. Fuhr,G., Hagedorn,R., MUller,T., Wagner,B. and Benecke,W. (1991) Proc. NicmElectro-Memanica~ System Ccng., Nara. 259-264 7. Wyman,J. and Mdh?kin,T.L.(1933)J. Am. Qlem. Soc. 55, 908-914 8. Greenstein,J.P., Wyman,J., and Cdm,E.J. (1935) t l . Rol. olern. Soc. 57, 637-642 9 . Ferry,J.D. and Wley,J.L. (1941) J. Am. Qlem. Soc. 63, 272-278 10. Srhwan,H.P., Srhwarz,G., Manuk,J. and Pauly,H. (1962) J. F@s. Qlern. 66,2626-2635 11. Muller,G. Van der Tauw,F. Zwolle,S. and Mande1,M. (1974) Biophys. chem. 2, 242-254 12. Hayakawa,R., Kanda,H., Sakrmot0.M and Wada,Y. (1975) Japnes Journal of Applied physics 14, 2039-2052 13. mlson,~,(1939) blloid Zeitschrift 68, 51
