Bloch D B, San Martin J E, Rauch S D, Moscicki R A, Bloch K J
Department of Medicine, Harvard Medical School, Boston, USA.
Arch Otolaryngol Head Neck Surg. 1995 Oct;121(10):1167-71. doi: 10.1001/archotol.1995.01890100075013.
To identify the 68-kd target of antibody in serum samples from patients with idiopathic, progressive, bilateral sensorineural hearing loss.
To purify target protein from renal extracts using gel filtration, ion-exchange chromatography, and polyacrylamide gel electrophoresis and to transfer to nitrocellulose membranes. The purified protein was digested with trypsin, and peptide fragments were separated by high-pressure liquid chromatography.
One fraction obtained by high-performance liquid chromatography contained a peptide of 2776 molecular weight. The sequence of a stretch of 22 amino acids within this peptide was identical to that of amino acids 424 through 445 of heat shock protein 70 (HSP70). On Western blotting, monoclonal antibody directed against HSP70 (but capable of recognizing both constitutive HSP70 [HSC70] and stress-inducible HSP70) reacted with the purified 68-kd protein. We compared the reactivity of serum samples from six patients with idiopathic, progressive, bilateral sensorineural hearing loss, as well as monoclonal antibody to HSC70, and monoclonal antibody to HSP70 with renal extract. The pattern obtained suggested that patient antibodies are preferentially directed at HSP70.
The target of antibody in serum samples from patients with idiopathic, progressive, bilateral sensorineural hearing loss is HSP70.
鉴定特发性、进行性双侧感音神经性听力损失患者血清样本中抗体的68-kd靶标。
使用凝胶过滤、离子交换色谱和聚丙烯酰胺凝胶电泳从肾脏提取物中纯化靶蛋白，并转移至硝酸纤维素膜上。将纯化后的蛋白用胰蛋白酶消化，肽片段通过高压液相色谱进行分离。
通过高效液相色谱获得的一个级分含有一个分子量为2776的肽段。该肽段内一段22个氨基酸的序列与热休克蛋白70（HSP70）的424至445位氨基酸序列相同。在蛋白质印迹法中，针对HSP70的单克隆抗体（但能够识别组成型HSP70 [HSC70]和应激诱导型HSP70）与纯化后的68-kd蛋白发生反应。我们比较了6例特发性、进行性双侧感音神经性听力损失患者的血清样本以及针对HSC70的单克隆抗体、针对HSP70的单克隆抗体与肾脏提取物的反应性。所得模式表明患者抗体优先针对HSP70。
特发性进行性双侧感音神经性听力损失患者血清样本中抗体的靶标是HSP70。
