PULSE POLAROGRAPHY AND INTRAVASCULAR OXYGEN ELECTRODES

Br. J. Anaesth. (1983), 55, 559

PULSE POLAROGRAPHY AND INTRAVASCULAR OXYGEN ELECTRODES I. L. SCOTT, A. M. S. BLACK, P. MAVNARD AND C. E. W. H A H N

Over the past two decades, a great deal of effort has been devoted to the quest for stable and accurate intravascular PO2 electrodes suitable for use in vivo. Many ingenious devices have been reviewed by Fatt (1976) and, in addition, the difficulties surrounding the physical principles, and the design, of these transducers have been considered by Rolfe (1976) and Hahn (1981). The transducers are almost all variations on the membrane-covered polarographic design of Clark (1956). As well as protecting the cathode from contamination, the membrane restricts the access of oxygen to the cathode. With an appropriate polarizing voltage applied to the cathode, a current is measured which is proportional to the PO2 in the "surface layer" of sample immediately adjacent to the outer surface of the membrane. A continuous polarizing voltage has been used in all instances (Hahn, 1981). The problem with the polarographic measurement of oxygen is that, of necessity, the electrode consumes the oxygen which it measures. This tends to deplete the oxygen in the surface layer of the sample and to cause the electrode to under-read. Unless the sample is well stirred, or is flowing rapidly past the electrode (to replenish the oxygen in the surface layer), this under-reading becomes appreciable. It is termed the "flow" or "oxygen consumption effect" (Rolfe, 1976). A thick impermeable membrane decreases the oxygen consumption, I. L. SCOTT, B.SC; A. M. S. BLACK, B.M., B.CH., M.A., D.PHIL., F.F.A.R.A.C.S.; P. MAYNARD, H.N.C; C. E. W. HAHN, M.A., B.SC.,

M.SC., D.PHIL.; Nuffield Department of Anaesthetics, Radcliffe Infirmary, Oxford, OX2 6HE. Correspondence to C.E. W.H.

but brings its own problems; the sensitivity tends to decrease and, since changes in PO2 take longer to be transmitted through the membrane to the cathode, the response time is prolonged. A thinner membrane increases the rapidity of response but also increases the oxygen consumption. Thus, each new design of in vivo electrode reflects a compromise between "response time" and "flow" (or "oxygen consumption effect"), but all within limits imposed by continuous polarization. "Pulse polarography" is another approach, in which the polarizing voltage is applied intermittently as a series of square-wave pulses. Davies and Brink (1942) demonstrated that this could eliminate the "flow-effect" since, although oxygen consumption still occurs, it is only a small fraction of what it would be under continuous polarization, and depletion of the oxygen in the surface layer of the sample tends not to occur. Several workers have applied pulse polarography to in vitro oxygen analysis (Kunze and Lubbers, 1973; Saulson, 1973; Zick, 1976), but it has not been tested in vivo until recently. We have used pulses of different voltages in commercial intravascular electrodes to obtain separate measurements of PO2 and P N 2 O from a single sensor (Brooks etal., 1978,1980; Hahn etal., 1979). However, commonly these commercial electrodes have a time to 90% response of about 60 s and as we required electrodes with a more rapid response time for certain physiological studies, further modifications were undertaken. Using a number of commercial electrodes, the thickness of the protective membrane was modified, and the findings on the relationships between elec© The Macmillan Press Ltd 1983

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The membranes covering standard polarographic m vwoPO2 electrodes were modified with a view to altering their response times. Their performance was examined under constant and pulsed polarization. Pulse polarization diminished the under-reading of the P02 signal which occurred at low flow rates. The stability of the electrode signal was greatly improved by integrating the electrode current over the pulse period. The combination of thin membranes and pulsed polarography will provide a P02 electrode with a fast time response without under-reading the true in vino PO2.

560

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trode current outputs, response time, and the susceptibility to flow effect are reported. In addition, we have compared two methods of processing the current response to a polarizing voltage pulse, with a view to improving the stability of the electrode.

Electrodes Searle (G. D. Searle and Co., size 5FG) intravascular electrodes were used throughout. The original membranes were carefully removed with a scalpel blade, and after the electrode tips had been completely cleaned, and coated with potassium chloride crystals, they were dipped into solutions of polystyrene in toluene. The viscosity of the solution was varied empirically so that a thick or thin film of liquid membrane would remain adherent to the tip of the electrode. These "dip-coated" membranes

teat electrode thermocouple

L electrode housing tuba* with water Jacket _ . Tuohy boret

tonometer with water jacket

wlndkeetel with water jacket

FIG. 1. Schematic diagram showing the in vitro circulation circuit for testing the intravascular electrodes. The arrows indicate the direction of flow.

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MATERIALS AND METHODS Electrode "test-rig" An in vitro test-rig was constructed (fig. 1) which consisted of two circuits, each with its own tonometer and "housing tube" for the electrode under test. The housing tube was straight, 31 cm long with an internal diameter of 6 mm. A peristaltic pump circulated 0.9% w/v saline through both circuits separately, at a flow rate of 0-360mlmin~'. This corresponded to a flow velocity down each electrode housing tube of 0-20 cm s~'. "Windkessel" compliances smoothed the pulsatility imparted by the pump. The whole apparatus was maintained at 37 °C by a water jacket, heater and thermostat. Although the two circuits were separate, a number of "crosspiece" connections existed, and were controlled by appropriately placed clamps. For the studies of "flow effect", only one circuit was used, and the saline therein was tonometered into equilibrium with 100% oxygen ("hyperoxic saline"). For the

response time studies, both circuits were used, the second tonometer being used to equilibrate the saline in the second circuit with pure nitrogen ("anoxic saline"). When an acute change of the saline in the housing tubes was required, from hyperoxic to anoxic or vice versa, the clamps on the interconnections were simultaneously moved onto the alternate limbs of the cross pieces. This effectively interchanged the housing tubes and their contained electrodes between the two circuits. A thermocouple was positioned adjacent to the tip of each electrode to confirm that the temperature did not change with changes in flow or with changes between hyperoxic and anoxic saline.

561

INTRAVASCULAR FO2 ELECTRODES were dried with a hot air blower. Not all of the modified electrodes functioned well. They were all tested for linearity before being included in the study.

Pulsing regimes and response processing

Two regimens of pulsing were examined. In one, the square wave pulse was "on" for 1 s and "off" for 6 s; in the other, it was "on" for 2 s and "off" for 5 s. In each case the pulse was repeated every 7 s. The current response of the electrode was converted by

Response time studies

Response times were studied with the electrode continuously polarized. Acute changes from hyperoxic to anoxic saline, and vice versa, were effected by interchanging the electrode housings as described above. (The apparatus was built to allow two electrodes to be tested simultaneously, but only one electrode was tested at a time in these studies.) The "hyperoxic-to-anoxic" and "anoxic-tohyperoxic" changes were both recorded on a Bryans

200nA electrode current output

r -650 mV polarizing voltage

2s

5s

FIG. 2. The electrode response to a polarizing voltage pulse. Bottom: polarizing voltage regimen. Top: current transient, showing the delay (100 ms) before integration. Shaded area is proportional to oxygen concentration.

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Linearity study The electrodes were inserted in random succession into Adams' tonometers containing saline equilibrated at 37 °C with pure oxygen, pure nitrogen or air. The gas bubbles kept the saline well stirred. The linearity of the readings was checked with the electrodes polarized continuously and by intermittent pulsing. For this and throughout the rest of the study, the polarizing voltage was —0.65 volts, with respect to a silver-silver chloride reference electrode. Three electrodes were discarded at this stage because of non-linearities under one or both conditions of polarization. Fifteen passed to the next stage of the study.

conventional means to a voltage, and this was passed through an analog-to-digital converter to a microcomputer (Research Machines, 38OZ). The pulse and response in a "2 on/5 off" regimen are illustrated in figure 2. The microcomputer processed the response in two different ways. The first was to measure the current amplitude at the end of the pulse (BC in figure 2). The second was to obtain the current - time integral (area ABCD in figure 2). These two estimates of the same PO2 were printed as a pair for each voltage pulse, using a Qume Sprint (5 series) printer. Since the estimate of PO2 was more stable using the current - time integral (see later), this became the method of choice in reporting the results of pulse polarography in these studies.

BRITISH JOURNAL OF ANAESTHESIA

562 X-Y-t recorder (Model 29000A3), used in the Y-t mode. The time courses were replotted by hand on semilog paper to obtain the time to 90% completion of the response, tK.

Electrode temperature coefficient During the studies of response time and "flow effect", the electrode temperature was maintained strictly at 37 °C. At the end of the flow studies, the heater was switched off while the pump continued to circulate the hyperoxic saline at the maximal velocity. For eight of the nine electrodes which entered the flow study, the decrease in electrode output, for each 0.1 °C decrease in temperature, was noted for the temperature range 37-36°C. THEORY Sensitivity/response time for continuous polarization If the tip of the measuring cathode presents a flat surface to the sample, and if oxygen can only approach from a direction perpendicular to the surface, the relationship between the electrode current and PO2 during continuous polarization is determined by the area of the surface (A), the thickness of the membrane (d), and the permeability of the membrane to oxygen ( P J : i OC 4a • p<* • p ° 2

1 S~2

r«, oc

A plot of f9o against S2 should give a rectangular hyperbola, and a plot of log 190 against log 5 should have a slope of - 0 . 5 . Current response to pulsed polarization For a pulse of polarizing voltage, the current response is a complex function of PO2 and the time (f) after the start of the pulse. The solubility ( O and diffusibility (D m ) of oxygen in the membrane are also determinants. For pulses of duration between 0.15 and 10 s (Mancy, Okun and Reilley, 1962; Hitchman, 1978; Hahn, 1980), the relation can be considerably simplified: i oc

. A

PO2

If the sample time (t) is kept constant, i is proportional to POj for a given electrode. The integral of i with respect to t (the current - time integral: fig. 2), will also be proportional to Po2. RESULTS AND DISCUSSION

Sensitivity and time to 90% response A plot of the square of electrode sensitivity (S 2 ) against (*> in seconds (fig. 3) appears at first sight to be a rectangular hyperbola, as expected. However, it appears to have an asymptote of 9 s on the {90 axis, which was not expected from the simple theory given above. The finding was confirmed by plotting tgo against the reciprocal of S 2 (not shown). The reasons for this finding are probably complex, including failure to bring about an accurate square wave change between hyperoxic and anoxic saline, and discrepancies between the reality of our modified electrodes and the simplified model behind the theoretical expectations. It must be acknowledged that, in reality, the electrode response time cannot

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"Flow effect" studies These were carried out after the response time studies, but only nine electrodes passed to this stage. All nine completed "flow effect" studies in the continuously polarized mode, but only five completed the whole of the planned experiment with both regimens of pulsing. The attrition rate probably reflected imperfections in our technique of "dipcoating". The circuit with hyperoxic saline was used, and the output of the peristaltic pump was varied to change the flow velocity past the electrode tip. Every second reading was taken at the maximal velocity of 20 cm s"1 to check for electrode drift, and the alternate readings were taken at velocities varying randomly between 8 and 10 values in the range 2 0 - 0 cm s*1. Sufficient time was allowed after each change in velocity to ensure that the electrode had reached its steady response. At each submaximal velocity, the output readings were referred (as percentages) to the mean of the readings for the maximal flow states immediately before and after.

If one defines the electrode sensitivity (5), as the electrode current per unit POj (5 =i/PO2 units nA kPa" 1 ), then S is proportional to (A.PaJd). After a step change in PO2, the electrode output should change in an exponential fashion (Hahn, 1980) such that the response to the change is 90% complete in a time (r^) which is proportional to (dV-Om), where Dm is the oxygen diffusion coefficient in the membrane material (Hahn, 1980,1981). From this one can deduce that t»is proportional to the reciprocal of S1 (inverse square law):

563

INTRAVASCULAR P(h ELECTRODES

_

6 4

TlO

9s

20

30

40

50

60

70

80

90

100 110

FIG. 3. A plot of the square of the electrode sensitivity (5)(nAkPa~') against the electrode time response (s) to 90% of the final signal, when the electrode was presented with a square wave challenge from nitrogen to 100% oxygen, or vice vena. The dotted line indicates a 9 s asymptote on the time axis (see text).

10.0

5.0 4.0 30 line of slope -0.5 2.0

2. <

1.0

0.5 0.4 0.3 0.2

0.1

2

3

4

5

10

20

30

40 50

100

f g O - 9 (s)

FIG. 4. A log/log plot of the electrode sensitivity against («so-9) s. A line of slope - 0 . 5 is drawn through the experimental points for comparison, but does not represent a line of best fit.

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'90

be completely defined by a monoexponential. This would be especially true for a dip-coated membrane which does not have a uniform thickness. More comprehensive explanations of the time-course of the electrode response have incorporated edge diffusion (Benedek and Heideger, 1970), non-uniformity of the membrane and electrolyte layers (Linek and Benes, 1977) and incomplete reduction of oxygen at the electrode surface (Hahn, Davis and Albery, 1975). Accepting an asymptote of 9 s as an empirical observation, plots of log (f«)-9) v. log 5 were performed to test whether the slope was - 0 . 5 (fig. 4). Although several lines of slope - 0 . 5 could be drawn through the points, the scatter of the points did not allow any conclusions as to which line, if any, was the most appropriate. This much is readily explicable. The theoretical expectations were based on the assumption that the thickness of the electrode m a n -

564

Processing of electrode output Before comparing the effects of flow in the face of continuous and pulsed polarization, the effects of the two different methods of assessing electrode output were investigated, to see whether the current-time integral gave a more stable reading than the single fixed-time current reading (see above). Previous publications (Brooks et al., 1978; 1980; Hahn et al., 1979) reported the latter method since the data capture utilized an analog sampleand-hold unit. The microcomputer used in this study could perform and compare both techniques. Figure 5 illustrates a comparison in a single electrode. With the maximal flow of hyperoxic saline past the electrode tip to provide a steady POj, the output of the electrode was charted for 20 consecutive pulses. The 20 fixed-time current readings and the 20 current-time integrals were all expressed as a percentage of their respective means, and plotted against time. The fixed-time method showed a much greater percentage variation about its mean than did the current - time integrals. Presumably "noise" or unidentified extraneous interference on the electrode signal affected the former, but had markedly less effect in the latter because of the integration over time. Although not all electrodes

were as "noisy" as that illustrated in figure 5, the current-time integral always improved on any instability seen with the fixed-time method. Thus, the integration method was used to express the results of the pulse polarography in the flow studies. Studies of "flow effect" Nine electrodes were studied for "flow effect" under continuous polarization. In theory, a greater decrease in electrode output with a decrease in flow was to be expected for those electrodes which also exhibited the greatest sensitivity and fastest response time. Such a relationship was not obvious, but the sample size was small and there may have been some imprecision in inducing the acute changes necessary for accurate measurements of response time, and the theory was probably oversimplified (see above). More importantly, in the five electrodes which survived the full regimen of pulse polarography, the "flow effect" was always diminished by pulse polarography and the 1-s pulse was clearly more effective than the 2-s pulse in three of the five electrodes (figs 6,7). For both continuous and pulsed polarization the signal at flow velocities less than 20cms" 1 are expressed as a percentage of the signal at 20 c m s " ' ( V^. The velocity axes on these plots are expanded with respect to the axes on the corresponding plots of other workers (Schuler and Kreuzer, 1967; Eberhard, Fehlmann and Mindt, 1979). The "flow effects" seem less abrupt, but the resolution with respect to flow is greater. Temperature coefficient Six of the eight electrodes for which temperature coefficients were studied had f^ less than 20 s and temperature coefficients of +3.3% per °C, SD 1% per °C. The remaining two electrodes with Un greater than 20 s had temperature coefficients of + 3.3% per °C and +2.8% per °C. Thus the modification which provides the benefit of a faster time response does not seem to entail the disadvantage of an appreciably greater temperature coefficient. CONCLUSIONS (1) It is feasible to modify the properties of commercial intravascular electrodes by modifying the protective membrane. (2) In the resulting electrodes, those with an adequately fast response time can be predicted from their output sensitivity to PO2. (3) The stability of the output signal from a pulsed

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branes could be altered without altering the permeability (Pn) and diffusibility (£>m) of oxygen in the membrane, and that the geometry of the electrodes was not altered. These conditions might not have been fulfilled because the thickness was altered by altering the composition and viscosity of the membrane solution into which the electrodes were dipped. In addition, there may well have been variations in the area of cathode (A ) between the different electrodes. Thus, the scatter of the points in figure 4 must be attributable in part to the fact that they are not several points on a single relationship, but are single points on several relationships. From a practical standpoint, the discrepancies between expectation and observation are not important. Figure 3 shows that, if the electrode sensitivity is greater than about 1 nAkPa" 1 , tx> is likely to be less than 20 s, whereas if S is less than 1 nA kPa" 1 , then fa, may be greater or much greater than 20 s. Thus, the studies have shown that the membrane properties of commercially available electrodes can be varied to produce electrodes with faster response times, and that the electrodes which have such a fast response time can be predicted from a simple measure of electrode sensitivity, S.

BRITISH JOURNAL OF ANAESTHESIA

INTRAVASCULAR P(h ELECTRODES

565 Electrode output • Area under current transient ° Height at the end of current transient

20 15 10 5 0 -5

r -15 Q -20

20

15

10 Number of pulses

FlG. 5. A comparison of the variation in the electrode output signal (under constant flow and oxygen concentration conditions) when displayed as the area under the current transient, or as the height at the end of the current transient. The ordinate axis is expressed as the difference of each electrode output signal from the mean signal over the recording time (140s, 20 pulses).

100 »-—

95

;

S

/

• o *-

i

i

i

8

12

16

A / E

Duration of pulse

'

• o *

1 s 2s continuous

¥

90

V RS

4

Flow v e l o c i t y

I max

20

v(cm

FIG. 6. Electrode output, expressed as a percentage of the maximum output, plotted against the velocity of oxygenated saline flowing past the electrode tip. The output is shown for the electrode either polarized continuously, or pulsed with 1 -s and 2-3 regimens. A clear improvement is seen between the 1-8 and 2-» pulse regimens; and between pulsing and continuous polarization.

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566

100 -

-^&*>~ Z-

o Duration of pulse

^^

mr

95 "

*

1^ A

*

ls continuous

90 --

1

1

1

1

8

12

16

20

Flow velocity l/lcnis' 1 )

FIG. 7. The electrode output plotted against flow velocity for another electrode, but this time there is not a clear improvement between the 1-s and 2-8 pulse regimens. Both pulse regimens are an improvement on continuous polarization.

electrode is improved by integrating the output over a period. (4) When under-reading of PO2 occurs at inadequate flow rates of sample past the electrode tip, this is diminished by using pulsed, rather than continuous polarization.

Fart, I. (1976). Polargraphic Oxygen Sensors. Cleveland: C. R. C. Press. Hahn, C. E. W. (1980). Techniques for measuring the partial pressures of gases in the blood. Part I—m vitro measurements. / . Phyt. E: Set. Instrum., 13, 470. (1981). Techniques for measuring the partial pressures of gases in the blood. Part II—in vivo measurements. / . Phys. E: Sci. Instrum., 14,783. Brooks, W. N., Albery, W. J., andRolfe,P.(1979). C^and ACKNOWLEDGEMENT N2O analysis with a single catheter electrode. An in vitro study. Mr I. L. Scon is supported by an M. R. C. Studentship Anaesthesia, 34, 263. (RS7914). Davis, A. H., and Albery, W. J. (1975). Electrochemical improvement of the performance of PO2 electrodes. Respir. Physiol.,25,109. REFERENCES Hitchman, M. L. (1978). Measurement of dissolved Oxygen. Geneva: John Wiley. Benedek, A. A., and Heidcger, W. J. (1970). Polarographic Kunze, K., and Lubbers, D. W. (1973). Absolute P02 measureoxygen analyser response: the effect of instrument lag in the ments with P02 electrodes applying polarising voltage pulsing; non-steady state reaeration test. Water Ret., 4,627. in Oxygen Transport to Tissue (eds H. I. Bicher and D. F. Brooks, W. N., Hahn, C. E. W., Foex, P., and Maynard, P. Bruley), p. 35. New York: Plenum Press. (1978). The simultaneous measurement of PO2 and PH2O in linek, V., and Benes, P. (1977). Multiregion, multilayer, vivo with a simple catheter electrode. Br. J. Anaesth., 50, nonuniform diffusion model of an oxygen electrode. Biotech1082P. nol. Bioeng., 19,741. Albcry, W.J. (1980). On-line R>2 and PN 2 O Mancy, K. H., Okun, D. A., and Reilley, C. N. (1962). A analysis with an in vivo catheter electrode. Br. J. Anatsth., 52, galvanic cell oxygen analyser. / . EUctroanal. Chem., 4,65. 715. Rolfe, P. (1976). Arterial oxygen measurements in the new born dark, L. C. (1956). Monitor and control of blood and tissue with intravajcular transducers; in /. E. E. Medical Electronics oxygen tension*. Trans. Am. Soc. Artif. Internal Organs, 2,41. Monographs, Vol. 18-22, Ch. 5. (edi D. W. Hill and B. W. Davies, P.W., and Brink, F. (1942). Microelectrodes for measurWatson). ing local oxygen tension in animals tissues. Rev. Sei. Instrum., Saulson, S. H. (1973). High speed pulsatile operation of minia13, 524. ture oxygen electrodes; in Advances m Experimental Medicine Eberhard, P., Fehlmann, W., and Mindt, W. (1979). An elecand Biology, (eds H. I. Bicher and D. F. Bruley), p.29. New trochemical sensor for continuous intravascular oxygen York: Plenum Press. monitoring. Bioulem. Patient Monitg., 6,16.

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567

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Schuler, R., and Kreuzcr, F. (1967). Rapid polarographic in vivo oxygen catheter electrodes. Rapir. PkysioL, 3, 90. Zick, G. L. (1976). Determination of oxygen tension by measurement of net charge transport. /. E. E. E. Tram. Biol. Eng., 23, 472.

ZUSAMMENFASSUNG

Die Mcmbranen von Oblkhen polarographischen in-vivo-POiElektroden wurden zum Zweck einer verfinderten Reaktionszeit modifiziert. Ihre Funkuon wurde unter konstanter und gepulster Polarographie gcpriift. Gepulste Polarographie veningerte das bei niedrigen Herzminutenvolumina auftretende Unterschatzen des PO2-SignaJs. Die Stabilitat des Elektrodensignals konnte durch Integrieren des Elektrodenstroms fiber die Pulsperiode stark verbessert werden. Die Kombination dunner Membranen mit gepulster Polarographie fuhrt zu einer P02-Elektrode mit schneller Reaktionszeit ohne Unterschatzung des tatsachlichen in-vivo-POi.

ELECTRODOS PARA POLAROGRAFIA DE IMPULSOS Y OXIGENO INTRAVASCULAR

RESUME

SUMARIO

Les membranes recouvrant des electrodes a Poi standard de polarographie in vivo ont etc modifiees dans le but d'alterer leurs temps de reponse. Leurs performances ont etc etudiees sous polflrisation cons tan te et pulsee. La polarisation pulsee diminnait la sous-estimation du signal Po2 qui surrient pour des debits bas. La stabilite du signal de l'electrode etait grandement amelioree par l'integration du courant de l'electrode sur la periode de pulsation. L'association de membranes fines et de la polarographie pulsee permettra 4c disposer d'une electrode a P02 pourvue d'une reponse rapide sans sous-estimation de la veritable P02 in vivo.

Se modifkaron las membranas que cubrian los electrodos normales de P02 para polarografia in vivo, con el fin de alterar sus tiempos de respuesta. Su funcionamiento se examin6 bajo polarizaci6n constante y pulsante. La polarizacion pulsante disminuyo la sublectura de la serial de P02 que tuvo lugar a regimenes de flujo bajo. La estabih'dad de la serial del electrodo mejoro considerablemente al integrar la corriente del electrodo a lo largo de todo el periodo del impulse La combinacion de membranas delgadas y de polarografia de impulsos provcera un electrodo de P02 con un tiempo de respuesta rapida y sin subleer in vivo el verdadero valor de P02.

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POLAROGRAPHIE PULSEE ET ELECTRODES INTRAVASCULAIRES A OXYGENE

GEPULSTE POLAROGRAPHIE UND INTRAVASKULARE SAUERSTOFFELEKTRODEN