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Keywords:

  • anaphylaxis;
  • drug allergy;
  • immunoglobulin E ;
  • immunoglobulin E antibody;
  • muscular blocking agent

Abstract

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

Background:  The general understanding is that a blood sample for analysis of immunoglobulin (Ig) E antibodies to an allergen suspected to cause an anaphylaxis cannot be drawn until several weeks after the reaction. As this is most unpractical, the changes in IgE antibody levels during anaphylaxis were studied to evaluate the possibility of using samples drawn at the time of the reaction.

Methods:  Immunoglobulin E antibodies to suxamethonium were quantitated with ImmunoCAP before, during and after an anaphylactic reaction occurring during anaesthesia using neuromuscular blocking agents.

Results:  Serum IgE antibody concentrations against suxamethonium in blood samples collected up to 6 h after the reaction were not different from those in samples drawn before or days and weeks after the anaphylaxis.

Conclusions:  A serum sample intended to trace the drug involved in an IgE-mediated anaphylactic reaction can be drawn in direct relation to the reaction.

In allergic anaphylaxis, the reaction is initiated by the causing agent reacting with its corresponding antibody or lymphocyte (1). Typical agents are classical allergens such as foods and hymenoptera venom (2) or, in the case of medical-related allergy, e.g. penicillin, neuromuscular blocking agents (NMBAs) and latex (3). Most cases of allergic anaphylaxis during anaesthesia are mediated by immunoglobulin (Ig) E antibodies but the use of dextran as a blood substitute can initiate an IgG antibody-mediated reaction (4).

Allergy to dextran was studied in detail in the late 1970s (5) and early 1980s and found to be mediated by immune complexes between IgG antibodies to dextran and dextran, which activate complement leading to an anaphylactic reaction (6). During the immune complex formation, most, if not all, serum IgG antibodies were consumed and thus a patient who had suffered such an anaphylaxis did not have measurable concentrations of IgG antibodies for several weeks, i.e. until significant amounts of new dextran IgG antibodies had been produced (5). Thus, a test for IgG antibodies to dextran could not be performed until 2–4 weeks after the reaction.

This knowledge of the neutralization and elimination of IgG antibodies in serum during an anaphylaxis seems to have been transferred to IgE antibodies, especially in severe, drug-induced, IgE-mediated reactions. Thus, a common understanding is that also IgE antibodies in serum are consumed during an anaphylactic reaction and therefore a serum sample for analysis of IgE antibodies to an allergen suspected for causing the reaction should not be drawn until several weeks after the reaction. Companies marketing tests for IgE antibodies recommend testing not before 2–3 weeks (7). However, a study investigating anaphylaxis to NMBAs (8) indicated that the drop in serum IgE antibody levels was rather small.

It would greatly simplify investigations of a suspected IgE-mediated anaphylaxis, if serum samples drawn in relation to the reaction could be used to identify the culprit. To evaluate the feasibility of this, a study of the kinetics of serum IgE antibody levels during an anaphylactic reaction was performed.

Material and methods

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

Subjects

Blood samples were collected from 15 patients, three men and 12 women, age range 20–60 years, with suspected anaphylaxis during general anaesthesia. Eight patients had elevated serum tryptase (>25 μg/l) and all had IgE antibodies (>0.1 kUA/l) to suxamethonium in relation to the reaction. Ten of the patients were atopic defined as an IgE sensitization to common allergens (1) detected as a positive PhadiatopTM test (Phadia AB, Uppsala, Sweden). All patients gave their informed consent allowing collection of the extra postreaction samples. The study was approved by the Regional Committee for Medical Research Ethics, Western Norway.

Trial design

Following the routine established by ‘The Norwegian Anaphylaxis Network’ (9) a blood sample should be drawn within 1–2 h after the incident. In this study, a sample was drawn after 10–345 min, median 45 min, denoted ‘per’, and a second sample on the day after the reaction (‘post’). A sample drawn prior to surgery (‘pre’), usually within a day or 2, is encouraged in the guideline. All samples were analysed for tryptase, IgE and IgE antibodies to suxamethonium and common, inhalant allergens. In addition, patients were asked to give two or three additional serum samples during the next 5–10 weeks after the reaction. To our knowledge, no individual was taking drugs for allergy during this period. The serum samples were kept frozen at −20°C pending analysis.

IgE and IgE antibodies

The serum concentrations of IgE (kU/l) and IgE antibodies (kUA/l) to suxamethonium (Rc202) and PhadiatopTM, a mixture of common, inhalant allergens [pollen from birch (t3), olive (t9), timothy (g6), mugwort (w6) and parietaria (w19), dander from cat (e1), horse (e3) and dog (e5), house dust mite Dermatophagoides pteronyssinus (d1) and the moulds Cladosporium (m2) and Alternaria (m6)], were determined by ImmunoCAPTM (Phadia AB) according to the manufacturer’s instructions using 0.1 kUA/l as the cut-off for a positive value.

Tryptase

Tryptase in serum was determined by a fluorescence immunoassay (ImmunoCAPTM, Phadia AB) following the manufacturer’s instructions but using 24 μg/l as the cut-off for a raised level, as established by the Haukeland University Hospital laboratory. The assay measures all extracellular varieties of α-tryptase and β-tryptase.

Statistical methods

Results are presented as median, geometric mean and range.

Results

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

The diagnosis of anaphylaxis to an NMBA was based on the clinical reaction supported by increased serum levels of tryptase and presence of IgE antibodies to suxamethonium in the ‘per’-sample. Eight of the 15 patients had a tryptase level above the cut-off, 24 μg/l, and an additional five patients had a value, which was more than twice as high as the concentration measured about 24 h after the reaction (‘post’-sample).

The 15 patients had a geometric mean serum IgE value of 466 kU/l (range: 39.2–6 700). Immunoglobulin E antibodies to suxamethonium, geometric mean 3.0 kUA/l (range: 0.5–15.6), were found in the ‘pre’-sample of all the 11 patients where such a serum sample was available. At the allergy follow-up, six of the eight investigated patients had a positive skin test to one or more NMBAs.

Serum concentrations of IgE antibodies to suxamethonium were measured before (11 patients), during and in most patients was followed for more than four weeks after the reaction (Fig. 1). The individual suxamethonium IgE antibody ratios of the per/pre samples were quite stable (geometric mean 1.0; range: 0.5–1.9; Table 1). None of the patients had nondetectable levels in the sample drawn at the time of the reaction. The same ratios for the samples post/per (geometric mean 1.2; range: 0.9–1.9; Table 1) indicated a small increase that in two of them (nos 1 and 11) stayed high for more than 2 weeks (Fig. 2).

image

Figure 1.  Changes in serum concentrations, kUA/l, of immunoglobulin E antibodies to suxamethonium. Time of sampling is presented, on a log-scale, in days from the reaction (‘per’) which was set to 1.0.

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Table 1.   Individual immunoglobulin E antibody concentrations, kUA/l, to suxamethonium and changes, expressed as concentration ratios, from before (‘pre’) to immediately after (‘per’) and to the day after (‘post’) the anaphylactic reaction
SamplePrePerPer/pre ratioPostPost/per ratio
17.814.71.920.61.4
20.50.51.00.51.0
3NT0.20.31.5
42.82.50.92.30.9
55.03.50.76.71.9
6NT0.30.51.7
7NT0.20.21.0
80.90.91.01.01.1
94.32.30.52.41.0
102.12.21.02.21.0
112.95.41.98.21.5
128.25.90.77.11.2
13NT3.44.61.4
1415.614.30.914.11.0
151.00.90.911.1
image

Figure 2.  Relative changes in serum concentrations of immunoglobulin E antibodies to suxamethonium. The level in sample drawn before the reaction, ‘pre’-sample, is set to 1. Time of sampling is presented, on a log-scale, in days from the reaction (‘per’) which was set to 1.0. Only patients where a ‘presample’ was available are included. Patients 1 (bsl00001bsl00001) and 11 (bsl00001- - -bsl00001) seem to have been boostered.

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The fluctuations seen in the suxamethonium IgE antibody concentrations during the clinical reaction, could, at least in part, be due to the intravenous fluids the patient received in relation to the clinical intervention. In an attempt to compensate for this factor, the suxamethonium IgE antibody values were calculated as a ratio to IgE, sometimes referred to as ‘total IgE’ (Fig. 3), and in the 10 Phadiatop-positive patients, also to the sum of IgE antibodies to common, inhalant allergens as measured by Phadiatop. These ratios showed essentially similar variations as the absolute concentration of suxamethonium IgE antibodies (data for Phadiatop not shown).

image

Figure 3.  The relative changes in serum concentrations of immunoglobulin (Ig) E antibodies to suxamethonium expressed in relation to the IgE concentration of the sample. Time of sampling is presented, on a log-scale, in days from the reaction (‘per’) which was set to 1.0. Only patients where a ‘presample’ was available are included.

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Discussion

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

Anaphylaxis to NMBAs during anaesthesia is a significant problem in some countries, e.g. Norway (10), France (11) and Australia (3). A centralized referral system for the diagnostic investigation has been available for some years and is now applied on a Nordic basis (12).

Allergic anaphylaxis is mostly caused by the interaction of an allergen with the corresponding allergen-specific IgE antibodies, which are present in serum and on mast cells and basophils of sensitized individuals (1). This shows that the patient has been exposed to the agent in question, or to another agent sharing the same allergenic epitopes, and developed the IgE sensitization.

During general anaesthesia, a large number of substances are administered to the patient. Within less than a minute the patient receives intravenously tens of milligrams of induction agents. In addition, there is exposure to antibiotics and latex. We hypothesize that under such circumstances, with even a moderate degree of IgE sensitization to some of these agents, a massive mast cell and basophil activation takes place (13). It is therefore nearly impossible to pinpoint the cause of the reaction without performing a follow-up investigation (14).

Tryptase is a neutral protease, mainly found in mast cells, and together with histamine it is a marker of mast cell activation (15, 16). An increased concentration, compared to the control sample, of serum tryptase, in close conjunction with the reaction, is a highly sensitive indicator of anaphylaxis, and the presence of an elevated serum tryptase has been reported to indicate an IgE-mediated mechanism of anaphylaxis during anaesthesia (17). In this study, eight patients had an increased level of tryptase. All the patients had IgE antibodies to suxamethonium, which was the criterion for inclusion in the study. For comparison, approximately 0.4% of Norwegian blood donors have low-to-moderate levels of IgE antibodies to suxamethonium (18).

The diagnosis of an IgE sensitization includes screening of a serum sample for IgE antibodies to allergens of interest (19) and samples from a patient investigated for anaphylaxis will, in most centres, be routinely tested. In the mid-1970s, it was found that during an anaphylactic reaction induced by infusion of dextran all IgG antibodies in serum could be consumed in immune complexes and thus disappear from circulation (5). Based on these findings and supported by some clinical observations (20, 21), it was recommended that a serum sample for analysis of IgE antibodies should not be drawn closer than a few weeks after a severe, IgE-mediated reaction (7). However, there are reports indicating that circulating IgE antibodies are not eliminated after severe hymenoptera venom reactions (22) and similar results were found in a study comprising 14 patients with grades 2 and 3 adverse reactions under anaesthesia (8). In the latter study, blood was sampled during, but not before, the reaction and at a follow up more than 8 weeks after the acute incident. The authors concluded that IgE antibodies towards NMBAs can be detected during the incident. This is in accordance with our findings. This approach will significantly simplify the diagnostic follow up after an anaphylactic reaction as serum samples for an IgE antibody diagnostic work-up could be drawn while the patient is still in the hospital.

In this study, IgE antibodies to suxamethonium were found before the anaphylactic reaction in all cases where a serum sample was available (11 patients), and in all patients after the reaction. Some fluctuations, including a moderate decrease compared to prereaction values, of serum IgE antibody levels to suxamethonium was seen, in some patients, in the sample drawn right after the reaction. Whether this is because of antibody consumption or dilution of the patients’ serum by infusion of plasma and blood volume expanders, is difficult to say. However, most importantly, in no case did the suxamethonium IgE antibodies become nondetectable. When the IgE antibody levels were presented as a ratio to IgE or, when present, IgE antibodies to common, inhalant allergens, these minor fluctuations were less prominent, although the IgE could partly result from plasma infusions during operation. This is probably a more relevant, patient-endogenous way to express the suxamethonium IgE antibody variations.

The most likely explanation for the difference between the disappearance from circulation of IgG antibodies to dextran and the very small effect on IgE antibodies is that the IgE-mediated reaction involves mast cell and basophil-bound IgE antibody molecules. Even if they represent a very small portion of the IgE antibody pool, they are the ones initiating the anaphylactic reaction. In contrast, the IgG antibodies to dextran will first bind the dextran in immune complexes, which then will activate complement, the inducer of the anaphylactic reaction.

It has been discussed whether an anaphylactic reaction to an NMBA has any significant booster effect on the IgE antibody levels to the major, NMBA allergenic determinant, the quaternary ammonium ion, QAI. All the patients in the present study were selected to have IgE antibodies to QAI at the time of the reaction. When the serum IgE antibody levels were followed up after the clinical incident, it seemed as if only two (nos 1 and 11) of the patients had a prominent suxamethonium IgE antibody booster increase compared to prereaction values, and an additional one (no. 15) a moderate, i.e. just <50%, response. In one of them (no. 1), also the anti-IgE/IgE ratio increased dramatically. Probably, the intense effect of the anaphylaxis on the circulation in most cases quickly eliminates the infused drug shortening the contact between the allergen and the immune competent cells. It is likely that repeated exposure over some time would be much more effective. However, daily exposure for 1 week of QAI IgE-sensitized healthy individuals to household chemicals and foods carrying the QAI did not result in an increased serum concentration of IgE antibodies to QAI, but might have delayed a normal decrease (23). This effect is in sharp contrast to the effect of a similar intake of a cough syrup with the active ingredient pholcodine, which stimulated IgE and IgE antibodies to QAI and the morphine allergenic epitope (23) in the order of 100-fold.

In conclusion, in anaphylaxis induced by NMBA in NMBA--IgE-sensitized individuals a small decrease of serum IgE antibodies to suxamethonium, which carries the representative allergenic epitope, QAI, was found in a few patients but in no case did the individual become serum-negative. Thus, a serum sample for analyses of IgE antibodies to QAI can be drawn right after the anaphylactic reaction. There is no reason to doubt that these findings do not apply to other drugs or allergens such as hymenoptera venom. Although an allergen booster increase during the 2–4 weeks follow up was seen in three of the patients, it is questionable if the marginally better diagnostic efficiency obtained by testing the patients again closely after the reaction would be cost-effective. Also, as the serum IgE antibody levels in many patients seem to decrease over a few months’ time a diagnostic re-visit cannot be too delayed. However, if a drug is suspected to be the cause of the anaphylaxis and no IgE antibodies could be detected in samples from the event, the tests should be repeated at the allergy work-up. Still to be elucidated is, if the prick test is reliable in the immediate time frame after an adverse drug reaction as antibody detection is only available for a limited number of drugs (24).

Acknowledgments

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References

The authors would like to thank Göran Nilsson, for evaluating statistical aspects and BMA Ingegerd Ågren-Andresson, MIAB, for performing the IgE and IgE antibody analyses.

References

  1. Top of page
  2. Abstract
  3. Material and methods
  4. Results
  5. Discussion
  6. Acknowledgments
  7. References
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