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Red cell binding of bilirubin
BRIEF
CLINICAL
LABORATORY
AND
OBSERVATIONS
Red cell binding of bilirubin Frank A. Oski, M.D., ~ and J. Lawrence Naiman, M.D. ~ BOSTON~
MASS.
Erythrocytes were obtained from normal adults with the use of ammonium-potassium oxalate, heparin, or acid-citrate dextrose solutions as anticoagulants. Cells were stored at 4 ~ C. and tested at intervals ranging from 1 hour to 1 week. T h e y were washed once with isotonic saline before use. Bilirubin determinations were performed in duplicate employing either the m e t h o d of Malloy and Evelyn or the spectrophotometric method of White, Abu Haider, and Reinhold? Red cell stroma was prepared in the manner described by Tishkoff, Robascheit-Robbins, and Whipple. 4 Washed erythrocytes were added to serum rich in bilirubin and mixed thoroughly. An aliquot of the mixture was removed immediately for determination of microhematocrit and serum bilirubin. T h e remainder of the mixture was then incubated at 37 ~ C. for 60 minutes in the dark with gentle agitation at 15 minute intervals. At the end of 60 minutes, bilirubin and micro hematocrit determinations were repeated and a Coombs test was performed on the mixture. Bilirubin determinations were carried out at 0 and 60 minutes on serum in a control tube incubated
T H E h u m a n red cell membrane contains appreciable amounts of lipid. 1 T h e ability of the red cell to absorb the unconjugated form of bilirubin, a lipid soluble compound, 2 was investigated, therefore. MATERIALS
AND METHODS
Serum rich in unconjugated bilirubin was obtained at the time of exchange transfusion from hyperbilirubinemic infants with erythrobtastosis fetalis.
From the Department of Pediatrics, Harvard Medical School and the Hematology Division o[ the Medical Service of The Children's Hospital Medical Center, Boston, Mass. Supported in part by Grants H-2405 and H T S - 5 2 5 5 [rom the Heart Institute of the National Institutes of Health, United States Public Health Service. e'Trainee in HematolofD, and Research Fellow in Pediatrics, Harvard Medical School. Address, Department of Pediatrics, Hospital o[ the University o[ Pennsylvania, Philadelphia 4, Pa. ~'~Research Fellow in Pediatrics, Harvard Medical School.
Articles submitted [or this section should not exceed 1,000 words and may contain 1 or 2 illustrations or tables. Bibliography should not exceed 7 references.
1034
Volume 63 Number 5
Brie/ clinical and laboratory observations
10 3 5
Table I. Bilirubin binding of Coombs' positive and Coombs' negative erythrocytes Bilirubin reduction per packed cell volume
Coombs' Coombs' Coombs' Coombs'
Cells negative positive negative positive
No. of experiments 19 14 10 2
Bilirubin method White et al White et al. Malloy and Evelyn Malloy and Evelyn
under identical conditions. Mixtures in which visible hemolysis occurred were excluded from the tabulations. Experiments employing red cell stroma or latex particles x" in place of erythrocytes were performed in a similar manner. At the time of exchange transfusion all blood removed was collected in a basin containing heparin. T h e cells and plasma were separated and their volumes measured. Aliquots of cells were removed, washed once in saline, and bilirubin was eluted with 5 or 10 per cent albumin solutions. After appropriate corrections were m a d e for the decrease in bilirubin due to dilution alone from the saline contained in the red cell suspensions (see formula in the Appendix) the bilirubin binding was calculated and expressed in terms of bilirubin binding per packed cell volume of 50 per cent. RESULTS
Erythrocytes demonstrated a consistent ability to absorb small amounts of bilirubin. T h e mean reduction for Coombs' negative erythrocytes employing the bilirubin method of White, Abu Haider, and Reinhold ~ was 1.59 mg. per cent per packed cell volume of 50 per cent compared to 0.75 mg. per cent per packed cell volume of 50 per cent for Coombs' positive erythrocytes (Table I ) . This difference in bilirubin binding between compatible and incompatible cells is significant (t = 3.2; p < 0.01). W h e n bilirubin determinations were performed by the ~Bacto-Latex 0.81, Difco Laboratories, Detroit, Michigan.
of Mean (rag. %) 1.59 0.75 1.97 0.3I
50% Range (mg. %) 0,55 to 3.10 0.00 to 1.70 1.00 to 2.60 0.00 to 0.62
% Reduction of initial bilirubin level per packed cell volume o[ 5O% Mean
Range
(%)
11.9 5.5 13.4 1.65
(%)
3.2 0.0 6.2 0.0
to 29 to 16 to 18.3 to 3.3
m e t h o d of Malloy and Evelyn, the magnitude of difference between Coombs' positive and Coombs' negative cells was even greater (Table I ) . T h e binding of bilirubin occurred almost instantly. Mixtures incubated for 1 hour rarely showed any further reduction of bilirubin levels. Neither storage for one week nor the anticoagulant adversely affected the ability of erythrocytes to bind bilirubin. Stroma prepared from compatible erythrocytes absorbed 0.014 mg. of bilirubin per milliliter of stroma while stroma from antibody-coated Coombs' positive erythrocytes did not absorb bilirubin. Latex particles also failed to lower the serum bilirubin concentration. W h e n cells previously incubated with bilirubin were suspended in albumin, significant amounts of bilirubin were eluted from the cells. A solution of 25 per cent albumin removed 0.052 mg. per milliliter; 10 per cent albumin, 0.030 mg. per milliliter; 5.0 per cent albumin, 0.037 mg. per milliliter; 1.0 per cent albumin, 0.019 mg. per milliliter. Saline suspensions removed only 0.012 rag. per milliliter. Erythrocytes removed at the time of exchange transfusion were coated with bilirubin. This bilirubin could be partially removed by albumin washing (Table I I ) . T h e amounts carried on the red cells were intermediate between those observed in vitro when sensitized and nonsensitized cells were compared. The cells removed at the time of exchange transfusion consisted of a . mixed population of sensitized and nonsensitized cells. Repeat exchange transfusions ap-
1 03 6
Brie[ clinical and laboratory observations
November 1963
Table II. Bilirubin binding of erythrocytes during exchange transfusion ~
Bilirubin removed By erythroeytes Exchange No.
By plasma I ConcenVolume tration (e.e.) (mg. %)
Total (rag.)
Total bilirubin removed (me.)
% o[ total removed by erythrocytes (%)
0.8 1.0 1.7 1.2 1.8 1.9 3.0 0.0 1.1 0.9 1.5 1.4 1.4
50.9 55.0 51.2 38.4 39.0 47.3 48.4 5.2 6.3 15.5 16.1 20.8 20.8
1.3 1.8 3.3 3.1 4.6 4.0 6.6 0.0 17.4 5.5 9.0 6.7 6.7
Con6en-
Total (me.)
Volume
(c.e.)
A.~ As A4 B1
279 306 300 264
17.9 18.0 16:5 14.3
50.1 54.0 49.5 37.2
201 204 215 176
B2
318
14.2
45.4
222
C1
235
2.2
5.2
140
D1
226
6.5
14.6
174
D~
235
8.3
19.4
140
tration in eluatet (me. %) 0.4 0.5 0.8 0.7 1.0 0.9 1.4 0.0 0.8 0.5 0.8 0.0 1.0
(5 %) (5%) (5%) (5%) ( 10% ) (5%) (10%) (5%) (10%) (5%) (10%) (5%) (10%)
~AI1 exchanges were performed for R h incompatibility. All cells were Coombs' positive. tFigures in parentheses refer to concentration of albumin solution used to elute bilirubin from erythrocytes.
peared to remove a greater percentage of bilirubin by virtue of erythrocyte bilirubin binding as would be expected when the population of sensitized cells had decreased. DISCUSSION Erthrocytes are capable of binding bilirubin. This ability is impaired when the erythrocyte is coated with antibody as demonstrated by a direct Coombs test. This binding ability appears to be a physicochemical property of the cell membrane in that it occurs almost instantly, stroma retains the ability to absorb bilirubin, and inert latex particles demonstrate no nonspecific adsorption properties. Albumin, a protein known to bind bilirubin, 3 appears to be capable of competing effectively with the lipid of the erythrocyte m e m b r a n e for bilirubin binding and removes some from its surface. Although this binding of bilirubin by erythrocytes is relatively small, it contributes to the reduction in bilirubin that occurs during exchange transfusions. A reduction of approximately 3.0 mg. per cent can occur for each 100 ml. of nonsensitized erythrocytes added to each 100 ml. of serum. This figure of 3.0 mg. per cent is in close agreement with the figure of 2.8 mg. per cent recently reported by Watson. 6 This slight but consistent
bilirubin lowering effect may have been responsible for the occasional observation of apparent benefit from simple transfusions in patients with hyperbilirubinemia due to erythroblastosis fetalis. Bilirubin absorbed on the red cell membrane may be a factor in the recently observed shortened red cell survivals noted in erythrocytes pre-incubated in solutions of unconjugated bilirubin. 7 Exchange transfusion replaces sensitized erythrocytes that have little capacity for bilirubin binding with erythrocytes that have greater binding potential. The bilirubin removed on erythrocytes should be calculated when determining the total a m o u n t of bilirubin removed at the time of exchange transfusion. We wish to express our appreciation to Mr. Dorlald McNamee and Miss Lorraine Kaneb for technical assistance. We are grateful to Dr. Sherwin Kevy for supplying us with the necessary patient materials, and to Dr. Louis K. Diamond for his guidance in this project. REFERENCES
1. Ponder, E.: Present concepts of the structure of the mammalian red cell, Blood 9: 227, 1954. 2. Overbeek, J. T. C., Vink, C. L. J., and Deen-
Volume 63 Number 5
3.
4. 5. 6. 7.
Brie[ clinical and laboratory observations
stra, H.: Solubility of bilirubin, Rec. tray. Chem. 74: 81, 1955. White, C., Abu Haider, G., and Reinhold, J. G.: Spectrophotometric measurement of bilirubin concentrations in the serum of the newborn by the use of a microeapillary, method, Clin. Chem. 4: 211, 1958. Tishkoff, G. I-I., Robascheit-Robbins, F. S., and Whipple, G. H.: Red cell stroma in dogs, Blood 8: 459, 1953. Martin, N. H.: Preparation and properties of serum and plasma proteins: Interactions with bilirubin, J. Am. Chem. Soc. 71: 1230, 1949. Watson, D.: The absorption of bilirubin by erythrocytes, Clin. Chim. Acta 7: 733, 1962. Sawitsky, A., Seifter, E., Bramson, S.: The toxicity of bilirubin on the mammalian erythrocyte, Proc. International Congress on Hematology, Mexico City, September, 1962.
10 3 7
APPENDIX Formula used to correct for fall in bilirubin concentration due to dilution alone from saline trapped in added red cell suspensions. V .... x Bl = amount of bilirubin reducV .... +V~,I. tion expected from dilution alone. V~,~. = Saline volume (volume of red cell mixture added-volume of packed cells) Vse,.. : serum volume Bl
: initial serum bilirubin concentration of red cell-serum mixture
Intracardiac fluid administration in shock John M. Verosky, M.D., * a n d I r a A. Budwig, Jr., M.D. EL
PASO,
TEXAS
S I N c E hypovolemic shock m a y be fatal, severe d e h y d r a t i o n in an infant with d i a r r h e a is an emergency situation. I n t r a v e n o u s fluids can usually be given through a superficial vein of the scalp or extremity or by c a n n u l a t ing a d e e p e r vein. Occasionally, however, technical difficulties resulting from vasoconstriction a n d vascular collapse m a k e p r o m p t intravenous t h e r a p y impossible. O t h e r routes, such as blind c a n n u l a t i o n of d e e p vessels or bone m a r r o w infusions, are available b u t require considerable experience. O n e of us has h a d experience with left h e a r t catheterization, a procedure involving only a small risk. T h e h a z a r d of h e m o p e r i c a r d i u m because of perforation of a superficial coronary vessel is real b u t limited. F o r the emergency administration of fluids, the right ventricle is probably the ideal t a r g e t ;
From the Medical Center, El Paso, Texas. ~Address, Medical Center, 1501 Arizona Avenue, El Paso 2, Texas.
it covers the greater p a r t of the anterior surface of the heart, its pressure is lower t h a n t h a t of the left ventricle so t h a t r e t r o g r a d e bleeding is less likely, the needle tract t h r o u g h the ventricular muscle is less likely to p e r m i t bleeding than the t h i n n e r wall of the atrium. T h e right a t r i u m is chiefly substernal, a n d attempts to p u n c t u r e its b o r d e r b r i n g one dangerously close to the a t r l o v e n t r i c u l a r groove, in which the right coronary artery lies. I t is not likely that there would be m a n y infants sufficiently ill to w a r r a n t this proced u r e a n d yet not have irreversible d a m a g e to the b r a i n or kidneys. F o r a n occasional infant, however, this m e t h o d is proposed as one which is easily p e r f o r m e d without previous extensive experience. C A S E REPORT A 3-month-old infant was hospitalized because of severe diarrhea. Weight was 10 pounds, 7 ounces; rectal temperature 109 ~ F.; pulse 280;
CLINICAL
LABORATORY
AND
OBSERVATIONS
Red cell binding of bilirubin Frank A. Oski, M.D., ~ and J. Lawrence Naiman, M.D. ~ BOSTON~
MASS.
Erythrocytes were obtained from normal adults with the use of ammonium-potassium oxalate, heparin, or acid-citrate dextrose solutions as anticoagulants. Cells were stored at 4 ~ C. and tested at intervals ranging from 1 hour to 1 week. T h e y were washed once with isotonic saline before use. Bilirubin determinations were performed in duplicate employing either the m e t h o d of Malloy and Evelyn or the spectrophotometric method of White, Abu Haider, and Reinhold? Red cell stroma was prepared in the manner described by Tishkoff, Robascheit-Robbins, and Whipple. 4 Washed erythrocytes were added to serum rich in bilirubin and mixed thoroughly. An aliquot of the mixture was removed immediately for determination of microhematocrit and serum bilirubin. T h e remainder of the mixture was then incubated at 37 ~ C. for 60 minutes in the dark with gentle agitation at 15 minute intervals. At the end of 60 minutes, bilirubin and micro hematocrit determinations were repeated and a Coombs test was performed on the mixture. Bilirubin determinations were carried out at 0 and 60 minutes on serum in a control tube incubated
T H E h u m a n red cell membrane contains appreciable amounts of lipid. 1 T h e ability of the red cell to absorb the unconjugated form of bilirubin, a lipid soluble compound, 2 was investigated, therefore. MATERIALS
AND METHODS
Serum rich in unconjugated bilirubin was obtained at the time of exchange transfusion from hyperbilirubinemic infants with erythrobtastosis fetalis.
From the Department of Pediatrics, Harvard Medical School and the Hematology Division o[ the Medical Service of The Children's Hospital Medical Center, Boston, Mass. Supported in part by Grants H-2405 and H T S - 5 2 5 5 [rom the Heart Institute of the National Institutes of Health, United States Public Health Service. e'Trainee in HematolofD, and Research Fellow in Pediatrics, Harvard Medical School. Address, Department of Pediatrics, Hospital o[ the University o[ Pennsylvania, Philadelphia 4, Pa. ~'~Research Fellow in Pediatrics, Harvard Medical School.
Articles submitted [or this section should not exceed 1,000 words and may contain 1 or 2 illustrations or tables. Bibliography should not exceed 7 references.
1034
Volume 63 Number 5
Brie/ clinical and laboratory observations
10 3 5
Table I. Bilirubin binding of Coombs' positive and Coombs' negative erythrocytes Bilirubin reduction per packed cell volume
Coombs' Coombs' Coombs' Coombs'
Cells negative positive negative positive
No. of experiments 19 14 10 2
Bilirubin method White et al White et al. Malloy and Evelyn Malloy and Evelyn
under identical conditions. Mixtures in which visible hemolysis occurred were excluded from the tabulations. Experiments employing red cell stroma or latex particles x" in place of erythrocytes were performed in a similar manner. At the time of exchange transfusion all blood removed was collected in a basin containing heparin. T h e cells and plasma were separated and their volumes measured. Aliquots of cells were removed, washed once in saline, and bilirubin was eluted with 5 or 10 per cent albumin solutions. After appropriate corrections were m a d e for the decrease in bilirubin due to dilution alone from the saline contained in the red cell suspensions (see formula in the Appendix) the bilirubin binding was calculated and expressed in terms of bilirubin binding per packed cell volume of 50 per cent. RESULTS
Erythrocytes demonstrated a consistent ability to absorb small amounts of bilirubin. T h e mean reduction for Coombs' negative erythrocytes employing the bilirubin method of White, Abu Haider, and Reinhold ~ was 1.59 mg. per cent per packed cell volume of 50 per cent compared to 0.75 mg. per cent per packed cell volume of 50 per cent for Coombs' positive erythrocytes (Table I ) . This difference in bilirubin binding between compatible and incompatible cells is significant (t = 3.2; p < 0.01). W h e n bilirubin determinations were performed by the ~Bacto-Latex 0.81, Difco Laboratories, Detroit, Michigan.
of Mean (rag. %) 1.59 0.75 1.97 0.3I
50% Range (mg. %) 0,55 to 3.10 0.00 to 1.70 1.00 to 2.60 0.00 to 0.62
% Reduction of initial bilirubin level per packed cell volume o[ 5O% Mean
Range
(%)
11.9 5.5 13.4 1.65
(%)
3.2 0.0 6.2 0.0
to 29 to 16 to 18.3 to 3.3
m e t h o d of Malloy and Evelyn, the magnitude of difference between Coombs' positive and Coombs' negative cells was even greater (Table I ) . T h e binding of bilirubin occurred almost instantly. Mixtures incubated for 1 hour rarely showed any further reduction of bilirubin levels. Neither storage for one week nor the anticoagulant adversely affected the ability of erythrocytes to bind bilirubin. Stroma prepared from compatible erythrocytes absorbed 0.014 mg. of bilirubin per milliliter of stroma while stroma from antibody-coated Coombs' positive erythrocytes did not absorb bilirubin. Latex particles also failed to lower the serum bilirubin concentration. W h e n cells previously incubated with bilirubin were suspended in albumin, significant amounts of bilirubin were eluted from the cells. A solution of 25 per cent albumin removed 0.052 mg. per milliliter; 10 per cent albumin, 0.030 mg. per milliliter; 5.0 per cent albumin, 0.037 mg. per milliliter; 1.0 per cent albumin, 0.019 mg. per milliliter. Saline suspensions removed only 0.012 rag. per milliliter. Erythrocytes removed at the time of exchange transfusion were coated with bilirubin. This bilirubin could be partially removed by albumin washing (Table I I ) . T h e amounts carried on the red cells were intermediate between those observed in vitro when sensitized and nonsensitized cells were compared. The cells removed at the time of exchange transfusion consisted of a . mixed population of sensitized and nonsensitized cells. Repeat exchange transfusions ap-
1 03 6
Brie[ clinical and laboratory observations
November 1963
Table II. Bilirubin binding of erythrocytes during exchange transfusion ~
Bilirubin removed By erythroeytes Exchange No.
By plasma I ConcenVolume tration (e.e.) (mg. %)
Total (rag.)
Total bilirubin removed (me.)
% o[ total removed by erythrocytes (%)
0.8 1.0 1.7 1.2 1.8 1.9 3.0 0.0 1.1 0.9 1.5 1.4 1.4
50.9 55.0 51.2 38.4 39.0 47.3 48.4 5.2 6.3 15.5 16.1 20.8 20.8
1.3 1.8 3.3 3.1 4.6 4.0 6.6 0.0 17.4 5.5 9.0 6.7 6.7
Con6en-
Total (me.)
Volume
(c.e.)
A.~ As A4 B1
279 306 300 264
17.9 18.0 16:5 14.3
50.1 54.0 49.5 37.2
201 204 215 176
B2
318
14.2
45.4
222
C1
235
2.2
5.2
140
D1
226
6.5
14.6
174
D~
235
8.3
19.4
140
tration in eluatet (me. %) 0.4 0.5 0.8 0.7 1.0 0.9 1.4 0.0 0.8 0.5 0.8 0.0 1.0
(5 %) (5%) (5%) (5%) ( 10% ) (5%) (10%) (5%) (10%) (5%) (10%) (5%) (10%)
~AI1 exchanges were performed for R h incompatibility. All cells were Coombs' positive. tFigures in parentheses refer to concentration of albumin solution used to elute bilirubin from erythrocytes.
peared to remove a greater percentage of bilirubin by virtue of erythrocyte bilirubin binding as would be expected when the population of sensitized cells had decreased. DISCUSSION Erthrocytes are capable of binding bilirubin. This ability is impaired when the erythrocyte is coated with antibody as demonstrated by a direct Coombs test. This binding ability appears to be a physicochemical property of the cell membrane in that it occurs almost instantly, stroma retains the ability to absorb bilirubin, and inert latex particles demonstrate no nonspecific adsorption properties. Albumin, a protein known to bind bilirubin, 3 appears to be capable of competing effectively with the lipid of the erythrocyte m e m b r a n e for bilirubin binding and removes some from its surface. Although this binding of bilirubin by erythrocytes is relatively small, it contributes to the reduction in bilirubin that occurs during exchange transfusions. A reduction of approximately 3.0 mg. per cent can occur for each 100 ml. of nonsensitized erythrocytes added to each 100 ml. of serum. This figure of 3.0 mg. per cent is in close agreement with the figure of 2.8 mg. per cent recently reported by Watson. 6 This slight but consistent
bilirubin lowering effect may have been responsible for the occasional observation of apparent benefit from simple transfusions in patients with hyperbilirubinemia due to erythroblastosis fetalis. Bilirubin absorbed on the red cell membrane may be a factor in the recently observed shortened red cell survivals noted in erythrocytes pre-incubated in solutions of unconjugated bilirubin. 7 Exchange transfusion replaces sensitized erythrocytes that have little capacity for bilirubin binding with erythrocytes that have greater binding potential. The bilirubin removed on erythrocytes should be calculated when determining the total a m o u n t of bilirubin removed at the time of exchange transfusion. We wish to express our appreciation to Mr. Dorlald McNamee and Miss Lorraine Kaneb for technical assistance. We are grateful to Dr. Sherwin Kevy for supplying us with the necessary patient materials, and to Dr. Louis K. Diamond for his guidance in this project. REFERENCES
1. Ponder, E.: Present concepts of the structure of the mammalian red cell, Blood 9: 227, 1954. 2. Overbeek, J. T. C., Vink, C. L. J., and Deen-
Volume 63 Number 5
3.
4. 5. 6. 7.
Brie[ clinical and laboratory observations
stra, H.: Solubility of bilirubin, Rec. tray. Chem. 74: 81, 1955. White, C., Abu Haider, G., and Reinhold, J. G.: Spectrophotometric measurement of bilirubin concentrations in the serum of the newborn by the use of a microeapillary, method, Clin. Chem. 4: 211, 1958. Tishkoff, G. I-I., Robascheit-Robbins, F. S., and Whipple, G. H.: Red cell stroma in dogs, Blood 8: 459, 1953. Martin, N. H.: Preparation and properties of serum and plasma proteins: Interactions with bilirubin, J. Am. Chem. Soc. 71: 1230, 1949. Watson, D.: The absorption of bilirubin by erythrocytes, Clin. Chim. Acta 7: 733, 1962. Sawitsky, A., Seifter, E., Bramson, S.: The toxicity of bilirubin on the mammalian erythrocyte, Proc. International Congress on Hematology, Mexico City, September, 1962.
10 3 7
APPENDIX Formula used to correct for fall in bilirubin concentration due to dilution alone from saline trapped in added red cell suspensions. V .... x Bl = amount of bilirubin reducV .... +V~,I. tion expected from dilution alone. V~,~. = Saline volume (volume of red cell mixture added-volume of packed cells) Vse,.. : serum volume Bl
: initial serum bilirubin concentration of red cell-serum mixture
Intracardiac fluid administration in shock John M. Verosky, M.D., * a n d I r a A. Budwig, Jr., M.D. EL
PASO,
TEXAS
S I N c E hypovolemic shock m a y be fatal, severe d e h y d r a t i o n in an infant with d i a r r h e a is an emergency situation. I n t r a v e n o u s fluids can usually be given through a superficial vein of the scalp or extremity or by c a n n u l a t ing a d e e p e r vein. Occasionally, however, technical difficulties resulting from vasoconstriction a n d vascular collapse m a k e p r o m p t intravenous t h e r a p y impossible. O t h e r routes, such as blind c a n n u l a t i o n of d e e p vessels or bone m a r r o w infusions, are available b u t require considerable experience. O n e of us has h a d experience with left h e a r t catheterization, a procedure involving only a small risk. T h e h a z a r d of h e m o p e r i c a r d i u m because of perforation of a superficial coronary vessel is real b u t limited. F o r the emergency administration of fluids, the right ventricle is probably the ideal t a r g e t ;
From the Medical Center, El Paso, Texas. ~Address, Medical Center, 1501 Arizona Avenue, El Paso 2, Texas.
it covers the greater p a r t of the anterior surface of the heart, its pressure is lower t h a n t h a t of the left ventricle so t h a t r e t r o g r a d e bleeding is less likely, the needle tract t h r o u g h the ventricular muscle is less likely to p e r m i t bleeding than the t h i n n e r wall of the atrium. T h e right a t r i u m is chiefly substernal, a n d attempts to p u n c t u r e its b o r d e r b r i n g one dangerously close to the a t r l o v e n t r i c u l a r groove, in which the right coronary artery lies. I t is not likely that there would be m a n y infants sufficiently ill to w a r r a n t this proced u r e a n d yet not have irreversible d a m a g e to the b r a i n or kidneys. F o r a n occasional infant, however, this m e t h o d is proposed as one which is easily p e r f o r m e d without previous extensive experience. C A S E REPORT A 3-month-old infant was hospitalized because of severe diarrhea. Weight was 10 pounds, 7 ounces; rectal temperature 109 ~ F.; pulse 280;