RATIONALE: We have previously defined IgE+ as the IgE on basophils from a subset of highly allergic, asthmatic subjects that release histamine after stimulation with histamine releasing factor (HRF), IL-3 and D2O. Since there have been reports of mouse monomeric IgE (termed HC IgE) causing mediator and cytokine release and phosphorylation (PY) events in rodent mast cells in the absence of antigen, we investigated if human IgE+ was HC IgE.METHODS: IgE+ was defined as causing > 10% histamine release (HR) to HRF after passive sensitization of dextran sedimented basophils. By definition IgE- did not support HR to HRF. Once defined, serum and various purified IgEs were used to stimulate highly purified basophils generated by double percoll gradients and negative selection and examined for ERK PY and cytokine production.RESULTS: 13 myeloma IgEs, 1 commercial IgE (Calbiochem) and gp120 IgE (Tanox) , all of which are IgE-, showed no ERK PY despite using 5 ug/ml. Serum from IgE+ individuals (5 ug/ml, n=2) and IgE- subjects (5 ug/ml, n=1) did show ERK PY, with more PY seen with IgE- serum. To further clarify this, IgE+ and IgE- were purified from the serum and demonstrated minimal ERK PY (5 ug/ml, n=2 respectively). The IgE in serum from 1 IgE+ and 1 IgE- donor caused IL-13 production from basophils.CONCLUSIONS: We have demonstrated ERK PY from 1 purified IgE+ and 1 purified IgE- subject. These IgE molecules must be further purified to assure they are monomeric. Thus, we conclude that IgE+ does not fully explain HC IgE. RATIONALE: We have previously defined IgE+ as the IgE on basophils from a subset of highly allergic, asthmatic subjects that release histamine after stimulation with histamine releasing factor (HRF), IL-3 and D2O. Since there have been reports of mouse monomeric IgE (termed HC IgE) causing mediator and cytokine release and phosphorylation (PY) events in rodent mast cells in the absence of antigen, we investigated if human IgE+ was HC IgE. METHODS: IgE+ was defined as causing > 10% histamine release (HR) to HRF after passive sensitization of dextran sedimented basophils. By definition IgE- did not support HR to HRF. Once defined, serum and various purified IgEs were used to stimulate highly purified basophils generated by double percoll gradients and negative selection and examined for ERK PY and cytokine production. RESULTS: 13 myeloma IgEs, 1 commercial IgE (Calbiochem) and gp120 IgE (Tanox) , all of which are IgE-, showed no ERK PY despite using 5 ug/ml. Serum from IgE+ individuals (5 ug/ml, n=2) and IgE- subjects (5 ug/ml, n=1) did show ERK PY, with more PY seen with IgE- serum. To further clarify this, IgE+ and IgE- were purified from the serum and demonstrated minimal ERK PY (5 ug/ml, n=2 respectively). The IgE in serum from 1 IgE+ and 1 IgE- donor caused IL-13 production from basophils. CONCLUSIONS: We have demonstrated ERK PY from 1 purified IgE+ and 1 purified IgE- subject. These IgE molecules must be further purified to assure they are monomeric. Thus, we conclude that IgE+ does not fully explain HC IgE.