| Prolactin (PRL) is a kind of polypeptide hormone synthesized by eosinocytes of anterior pituitary. Three fractions of immunoreactive PRL of different molecular weights are confirmed using gel filtration chromatography (GFC), they are 23 kDa monomeric PRL,45-60 kDa big PRL, and>100kDa big-big PRL or macroprolactin (MP). In normal serum, monomeric prolactin accounts for 85%-95% of the prolactin present and big-big prolactin, or macroprolactin, accounts for<1% of total prolactin. Hyperprolactinemia(HPRL) is a state of rising of serum PRL in female≥25ng/ml or in man PRL≥20ng/ml. The accurate quantitative analysis of prolactin is very important to clinic diagnosis, therapy, and prognosis. The existence of macroprolactin is one of the main reasons leading to misdiagnosis of hyperprolactinemia. It is becoming more and more important how to eliminate the interference of macroprolactin and enhance the veracity of hyperprolactinemia diagnosis.Gel filtration chromatography (GFC), the gold standard for quantifying bioactive monomeric prolactin in sera, is costly and labor-intensive.Where screening takes place, laboratories generally use polyethylene glycol (PEG) precipitation to differentiate macroprolactinemia from true hyperprolactinimia. While the feasibility of PEG precipitation on screening of hyperprolactinimia is still not be confirmed.Objective1. We established the method of GFC to separate three fractions of PRL in serum and investigated the basic conditions and procedures of this assay. We also compared the PEG precipitation to GFC method, finally, detected the feasibility of PEG precipitation on true hyperprolactinemia diagnosis.2. We used PEG precipitation to pretreat samples, and then assessed the morbidity of macroprolactinemia, so as to avoid misdiagnosis of hyperprolactinimia, and direct clinical diagnosis and therapy of this disease.Methods 1.26 samples showing high serum prolatin concentration were examined by BayerACS180 immunoassay. Each sample was divided into two halves, one half for GFC assay and the other for PEG precipitation. All those 26 samples were divided into two groups, ture hyperprolactinemia and macroprolacinemia, by GFC method. We detected the correlation between PEG precipitation and GFC method.2.200 cases of hyperprolactinemia were chosen and classified into two groups, according to their clinical status. GroupA included subjects with pituitary lesions hyperprolactinemia. GroupB includeed subjects with functional hyperprolactinemia. We also set a control group as group C including 100 healthy persons with regular menses, some previous pregnancies, drug free and without endocrinol disease. Serum prolactin concentration was measured befor and after PEG precipitation by automated chemiluminescencd system. Following the criterion of a PRL recovery≤40% was indicative of significant macroprolactinemia.Results1. The correlation between GFC and PEG pretreatment on determination of macroprolactinemia1.1 The prolactin concentration in serum samples of those three groups was measured before and after PEG precipitation. The levels of PRL of true hyperprolactinemia group and MP group before PEG pretreatment were 95.60ng/ml and 64.43ng/ml, no significant difference was found(P=0.113). And for the corresponding groups after PEG pretreatment, the levels of PRL were 66.86ng/ml and 12.00ng/ml, respectively., There was a significant difference between these two groups(P=0.000).1.2 The levels of monomeric PRL in serum were compared between the true hyperprolactinemia groups and macroprolactinemia groups using GFC or PEG pretreatment. There were no significant differences between the levels of serum monomeric PRL in the true hyperprolactinemia groups or the macroprolactinemia groups determined by those two methods. They were 68.96ng/ml vs 66.86ng/ml for the true hyperprolactinemia group(P=0.075), and 11.82ng/ml vs 12.00ng/ml(P=0.281) for the macroprolactinemia group.1.3 The recovery of monomeric PRL in serum were compared between the true hyperprolactinemia groups and macroprolactinemia groups using GFC and PEG pretreatment.The recoverys of serum monomeric PRL were 69.17% vs 76.93%,(P=0.052) in the true hyperprolactinemia groups determined by two methods; and the recoverys of serum monomeric PRL were 24.94% vs 19.89%(P=0.100) in the macroprolactinemia groups. No significant differences were found.1.4 There was a good correlation between results of PEG pretreatment group and GFC group. (r=0.844,P=0.000).1.5 Method imprecision of PEG pretreatment:PEG pretreatment in high value mixed serum yielded a within-CV 6.34% and totalCV 8.08%.The corresponding values for the low value mixed serum were 6.12% and 9.19%.2. Morbidity of macroprolactinemia in patients with hyperprolactinemia2.1 The levels of serum prolactin and recoverys were compared between pituitary lesions hyperprolactinemia, functional hyperprolactinemia, and the normal control group with and without PEG treatment.Serum prolactin levels of three groups (A,B,C) without PEG pretreatment were 88.17ng/ml,42.93ng/ml and 8.65ng/ml respectively, a significant difference was found.Serum prolactin levels of three groups with PEG pretreatment were 62.17 ng/ml,21.20ng/ml and 6.10ng/ml respectively, a significant difference was found.The PRL level of pituitary lesions hyperprolactinemia group was significantly higher than that of functional hyperprolactinemia group and normal control group not only before but also after PEG pretreatment(P=0.000), and thing is the same on hyperprolactinemia group and normal control(P=0.000).The serum prolactin recoverys within each group with PEG pretreatment show significant differences. The recoverys were 61.91%,50.11% and 69.50% respectively. The serum prolactin recovery was higher(P=0.000) in normal control than that of the other two groups (P=0.000), and the recovery of pituitary lesions hyperprolactinemia group was higher than that of functional hyperprolactinemia group(P=0.000).2.2 The prevalence of macroprolactinemia was 3%,25%,1% in group A,B,C,respectively.The presence of macroprolactin was correlated with the prevalence of PRL, especial in subjects with functional hyperprolactinemia. These results showed that the incidence of macroprolactin was higher (P=0.000) in group B than that of groupA (P=0.000) and group C (P=0.000).Whereas, there was no significant difference between group A and group C (P=0.312).Conclusion1. PEG pretreatment yielded results of monomeric PRL that correlated well with GFC (r=0.844,P=0.000) and was recommended as a routine screening of hyperprolactinemic sera for macroprolactin.2. The presence of macroprolactin is correlated with the prevalence of PRL, especial in subjects with functional hyperprolactinemia. The study showed that macroprolactin level was a significant factor for the misdiagnosis, unnecessary investigation, and inappropriate treatment. The PEG immunopreciptation procedure may be of particular importance to eliminate macroprolactin interference in PRL immunoassays, and enhance the diagnosis of hyperprolactinemia and follow-up. |
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