Immunology 1978 35 567
The effect of passive immunization against tetanus toxoid in man
MARIA PLASSARA, CATHERINE MANDALENAKI, E. PAPATHANASSIOU, EFTHYMIA VRACHNOU & P. ATHANASSIADES Department of Biology, Nuclear Research Center Demokritos, Aghia Paraskevi Attikis, Greece, 1st Department of Pediatrics University of Athens, and Department of Clinical Therapeutics, University of Athens, Greece
Received 5 January 1978; acceptedfbr publication 23 January 1978
Summary. The present study deals with the immunological response in man after passive immunization against tetanus toxoid. Treatment of man with equine antitetanus serum stimulates a rise of E rosette-forming T lymphocytes. The level of IgG also rises while the level of IgM falls. It is assumed that the foreign globulin acts as an antigen that evokes the T lymphocytes' co-operation for the humoral immune response with a switch from IgM to IgG synthesis.
effects of such serum on the leucocytes, the E rosette-forming T lymphocytes and the major immunoglobulin classes were studied. We found that man responds to the administration of tetanus antitoxoid by a rise in the number of lymphocytes and E rosette-forming T lymphocytes. The IgG level rises too, whereas that of IgM falls. MATERIALS AND METHODS Eleven healthy adult males aged 18-23 years were given an intramuscular injection of 1 ml (1500 iu) tetanus antitoxoid (Instituto Sieroterapico Milanese Serafino Belfanti). None of them had previously received tetanus antitoxoid immunization or any other passive immunization. Blood samples for white cell count, differential count, T-cell determination and immunoglobulin measurement were taken prior to and on the 5th and 12th day after administration of antitetanus serum.
INTRODUCTION
The prevention of tetanus in the wounded includes the use of tetanus antitoxin, the beneficial, effect of which in conferring passive protection is indisputable. Observations concerning the immune response in man after the administration of the antitetanus serum have dealt mainly with the kinetics of disappearance of the foreign protein and with hypersensitivity reactions. In the present study we have examined the immunological response in man after the administration of horse antitetanus serum. For this purpose the
Test for E rosette-forming lymphocytes 4 ml of heparinized blood were layered on top of 2 ml of 9% Ficoll-34 % Hypaque after the method of Harris & Ukaejiofo (1969). The purified lymphocytes were washed in Hanks's solution and adjusted to a concentration of 4 x 106/ml. Defibrinated sheep red blood cells (SRBC) were stored in Alsever's solution until used.
Correspondence: Dr Maria Plassara, Nuclear Research Center Demokritos, Department of Biology, Aghia Paraskevi Attikis, Greece. 0019-2805/78/0900-0567 $02.00 © 1978 Blackwell Scientific Publications
567
Maria Plassara et al.
568
They were then washed three times in normal saline and adjusted to a concentration of 30 x 106/ml in Hanks's solution. Inactivated foetal calf serum (FCS) was absorbed with SRBC at 370 for 1 h and subsequently with a mixture of human group A and B red blood cells at 40 for I h. A volume of 0-25 ml purified lymphocyte suspension was added to 0 25 ml FCS and left at room temperature for 1 h and then 0-25 ml SRBC suspension was added. After 10 min incubation at 370 the mixed cell suspension was centrifuged at 200 g for 5 min and kept 90
(a)
80
at 40 overnight. The sedimented cells were gently resuspended and rosettes were counted in a haematocytometer. At least 300 lymphocytes were scanned and lymphocytes binding five or more red blood cells were considered as rosette-forming cells. The percentage of rosette-forming T lymphocytes was calculated, and the absolute count of peripheral blood T lymphocytes was determined from the total lymphocyte count in the blood sample studied.
Estimation of serum immunoglobulin Serum IgG, IgM and IgA were measured by the radial immunodiffusion technique after the method of Mancini, Carbonara & Heremans (1965) using goat anti-human IgG, IgM and IgA. The statistical analysis was performed by the paired 't'-test.
RESULTS
5000
Following tetanus antitoxoid injection a rapid rise of E-rosetting cells was noted on the 5th day (Fig. 1). Prior to the tetanus antitoxoid administration the mean value of E-rosettes was 1340 per mm3 of peripheral blood or 49 % of total lymphocytes, whereas on the 5th day it rose to 2234 or 66%. This rise is statistically significant (P
The effect of passive immunization against tetanus toxoid in man
MARIA PLASSARA, CATHERINE MANDALENAKI, E. PAPATHANASSIOU, EFTHYMIA VRACHNOU & P. ATHANASSIADES Department of Biology, Nuclear Research Center Demokritos, Aghia Paraskevi Attikis, Greece, 1st Department of Pediatrics University of Athens, and Department of Clinical Therapeutics, University of Athens, Greece
Received 5 January 1978; acceptedfbr publication 23 January 1978
Summary. The present study deals with the immunological response in man after passive immunization against tetanus toxoid. Treatment of man with equine antitetanus serum stimulates a rise of E rosette-forming T lymphocytes. The level of IgG also rises while the level of IgM falls. It is assumed that the foreign globulin acts as an antigen that evokes the T lymphocytes' co-operation for the humoral immune response with a switch from IgM to IgG synthesis.
effects of such serum on the leucocytes, the E rosette-forming T lymphocytes and the major immunoglobulin classes were studied. We found that man responds to the administration of tetanus antitoxoid by a rise in the number of lymphocytes and E rosette-forming T lymphocytes. The IgG level rises too, whereas that of IgM falls. MATERIALS AND METHODS Eleven healthy adult males aged 18-23 years were given an intramuscular injection of 1 ml (1500 iu) tetanus antitoxoid (Instituto Sieroterapico Milanese Serafino Belfanti). None of them had previously received tetanus antitoxoid immunization or any other passive immunization. Blood samples for white cell count, differential count, T-cell determination and immunoglobulin measurement were taken prior to and on the 5th and 12th day after administration of antitetanus serum.
INTRODUCTION
The prevention of tetanus in the wounded includes the use of tetanus antitoxin, the beneficial, effect of which in conferring passive protection is indisputable. Observations concerning the immune response in man after the administration of the antitetanus serum have dealt mainly with the kinetics of disappearance of the foreign protein and with hypersensitivity reactions. In the present study we have examined the immunological response in man after the administration of horse antitetanus serum. For this purpose the
Test for E rosette-forming lymphocytes 4 ml of heparinized blood were layered on top of 2 ml of 9% Ficoll-34 % Hypaque after the method of Harris & Ukaejiofo (1969). The purified lymphocytes were washed in Hanks's solution and adjusted to a concentration of 4 x 106/ml. Defibrinated sheep red blood cells (SRBC) were stored in Alsever's solution until used.
Correspondence: Dr Maria Plassara, Nuclear Research Center Demokritos, Department of Biology, Aghia Paraskevi Attikis, Greece. 0019-2805/78/0900-0567 $02.00 © 1978 Blackwell Scientific Publications
567
Maria Plassara et al.
568
They were then washed three times in normal saline and adjusted to a concentration of 30 x 106/ml in Hanks's solution. Inactivated foetal calf serum (FCS) was absorbed with SRBC at 370 for 1 h and subsequently with a mixture of human group A and B red blood cells at 40 for I h. A volume of 0-25 ml purified lymphocyte suspension was added to 0 25 ml FCS and left at room temperature for 1 h and then 0-25 ml SRBC suspension was added. After 10 min incubation at 370 the mixed cell suspension was centrifuged at 200 g for 5 min and kept 90
(a)
80
at 40 overnight. The sedimented cells were gently resuspended and rosettes were counted in a haematocytometer. At least 300 lymphocytes were scanned and lymphocytes binding five or more red blood cells were considered as rosette-forming cells. The percentage of rosette-forming T lymphocytes was calculated, and the absolute count of peripheral blood T lymphocytes was determined from the total lymphocyte count in the blood sample studied.
Estimation of serum immunoglobulin Serum IgG, IgM and IgA were measured by the radial immunodiffusion technique after the method of Mancini, Carbonara & Heremans (1965) using goat anti-human IgG, IgM and IgA. The statistical analysis was performed by the paired 't'-test.
RESULTS
5000
Following tetanus antitoxoid injection a rapid rise of E-rosetting cells was noted on the 5th day (Fig. 1). Prior to the tetanus antitoxoid administration the mean value of E-rosettes was 1340 per mm3 of peripheral blood or 49 % of total lymphocytes, whereas on the 5th day it rose to 2234 or 66%. This rise is statistically significant (P
