Microelectrophoresis using small volumes of buffer

CIINICA

MICROELECTROPHORESIS

CHIMICA ACTA

USING

537

SMALL

VOLUMES

OF BUFFER

J. FISCHL Department,

Biochemical

“Assaf

Harofe”

Govevnment Hospital, Zerifivl (Isvael)

(Received January 8th, 1962)

In “micro” taken

electrophoresis,

little

for the run, for “macro”

features

of “micro”

electrophoresis

the low cost and simplicity cedures

for routine

PORT et al.1 in methods type

with

is gained

methods

by a reduction

also employ

are the reduction

of construction,

electrophoresis.

of apparatus,

acetate

Such an apparatus

in which

widely

as the supporting

the advantages

samples. simple

used.

KOHN’S~ would

by

pro-

RAPPA-

electrophoresis

however,

of this material

apparatus,

and reliable

was constructed

medium,

of serum

The important

in the size of the

and the rapid,

1957 and has since become

cellulose

of the amount

minute

suggested be fully

a new

realized.

CONSTRUCTION OF APPARATUS For height

the buffer

vessels,

2.0 cm are used.

two

small

Empty

plastic

boxes

Fig. I. Joining the cover-glass The

boxes

vessels two

thick

nylon

and about the

are glued

threads

are then

the perspex

with

are fixed

base

areas

cover

into

4 perspex

glued

to it (Fig. walls.

of 2.5 cm2 and

glasses

can be used.

boxes for the base of the apparatus.

I). To

the rim;

at the inner

with

x zz-mm

the lid of one of the boxes

of 2.5 cm (Fig.

4 mm below

assembly

through

together

by a distance

boxes

for zz

keep

the cellulose

each box plates

4 mm

from

strip

platinum

are bent

electrodes

in such a way

the two in place,

the outer

3.5 cm high and exactly

2). Two They

about

separating

acetate

edge

surrounding are inserted that

their

tip

J. I:lSCHI.

53s

Fig. 2. The apparatus

two

lntroduw 0 ml ol \-cxronal buffer vwscls. Cut :I 2.0 X rr.o-cm strip

l~and), moisten kvith the same buffer and insert the ends into the nylon

ready

for USC.

ionic strctngth 0.05) into each of that of cellulose xwtatt paper (do not touch by

(pH S.6;

for about 5 min, blot bet\vccn filter paper strips threads in the two compartments. An arch is

formed and, owing to the rigidity of the cellulose ncctate paper, tlic bridge supports itself in the air, with only the ends touching the walls of the: vcsscl. Apply about 5-S ,~ugof serum (or Hb hcmolysatej at the apex of the arch (i.e. in thtt middlo of the strip) by means of a capillary pipette or any othrr suitabk de\-kc. Cover the apparatus \vith n glass plate, and apply the currcnt. (;ood resolution can bo obtained in 30 min at 100 I,‘, in z h at ho V, and in g 11 at rq 1’ (Fig. 3). * Staining can be carried out by any convenient method, preferably with ligllt grwn or Ponceau 5 stains, and the evaluation mctry or by visual observa lion.

‘Ilie

chkf

xlvantagc~s

of the

micro

can bv performed

elcctrophorctsis

by clution,

apparatus

are

denliito-

tlit:

rapid

resolution of the c:lectrophcrogr;lm at 100 \- so that rtwlts can lx: obtained in 30 min, and the, lon \-oltage resolution, e.g. at 14 \-, which allows the use of dry bxtteric~ instcacl of ;I central paver supply. An clcctrophorc~is cxn thus hc performed in rcrnotc plnces I\:hcrc no clcctricity is n~~ailabl~~. Rot11 of thew ntlvantages are dlw to thcx !+mall size: of the: clcctrodc vcsscls and to ihc minimal amounts of buffer employed. ‘1’11euse of the apparatus is also very economical. Although ordinary filter paper- (suitalA: ior clcctrophoresk) can be used \\.itll t11c: becomes c~scwsive, considcrillg the> small nmount of sainplv, apparatus, “tailing”

MICROELECTROPHORESIS

539

and cellulose acetate is much better fitted for the purpose. The products from “0x0” and Schleicher and Schiill are suitable. Agar gel electrophoresis, with microscope slides as support, can also be performed, the time of resolution again being reduced. Thus,

agar immuno-electrophoresis is possible. Starch gel or block electrophoresis has not yet been tried.

Fig. 3. Electropherograms:

(I) IO min at 4oo V, (2) 30 min at 160 V, (3) 100 min at 80 V, (4) 120 min at 60 V, (5) 7 h at 14 V. In each case a normal (upper) and a myeloma serum (lower) was run on the same strip. (Note the faster moving M’ component.) Distance of migration of No. I = 160 mm and of No. 4 = 440 mm. ACKNOWLEDGEMENT

I express my thanks to Dr. H. STEGEMANN and his staff of the Max Planck Institute (Gbttingen) for their help in the experimental stage of this work, at the Symposium on Protein Analysis at Gottingen (1961). SUMMARY

The construction and use of a simple, economical micro electrophoresis apparatus is described in detail. With cellulose acetate paper as the supporting medium, electrophoresis requires 30 min. Dry batteries may be used as the power supply; at 14 V resolution is obtained in 5-7 h. REFERENCES 1 F. RAPPAPORT, F. EINHORN ’ J. KOHN, CZim. Chim. Acta,

AND J. FISCHL, Clin. Chim. 2 (1957)

Acta,

2 (1957) 390.

297. Clin. Chim.

Acta,

, (1962)

53,-x39