Cryz S J, Fürer E, Que J U
Swiss Serum and Vaccine Institute, Bern, Switzerland.
Infect Immun. 1991 Jan;59(1):45-50. doi: 10.1128/iai.59.1.45-50.1991.
Alginate from Pseudomonas aeruginosa 3064 was depolymerized by controlled heating in dilute acid. The resulting depolymerized alginate (Mr less than 60,000) was covalently coupled to toxin A with adipic acid dihydrazide as a spacer molecule and carbodiimide as a linker. The resulting conjugate was composed of toxin A and depolymerized alginate at a ratio of 4:1 and possessed an Mr of 260,000. The conjugate was nontoxic and nonpyrogenic. While native alginate (Mr greater than 640,000) given in a range of doses was poorly immunogenic in mice and rabbits, the conjugate induced high levels of antibody which bound to native alginate. Rabbits, but not mice, also produced an antitoxin immunoglobulin antibody response. Alginate derived from three other strains of P. aeruginosa competed with the homologous 3064 alginate for binding to anticonjugate antibody. This indicates that the conjugate elicits an antibody response able to recognize heterologous alginates. The serum from rabbits immunized with the conjugate was effective at promoting the uptake and killing of mucoid strains of P. aeruginosa by human polymorphonuclear leukocytes. In contrast, immunization with native alginate did not engender an opsonic antibody response. Rabbit anticonjugate antibody also neutralized the cytotoxic potential of toxin A.
通过在稀酸中控制加热，使铜绿假单胞菌3064产生的藻酸盐解聚。将所得的解聚藻酸盐（分子量小于60,000）以己二酸二酰肼作为间隔分子、碳二亚胺作为连接剂与毒素A共价偶联。所得的偶联物由毒素A和解聚藻酸盐以4:1的比例组成，分子量为260,000。该偶联物无毒且无致热原性。虽然在一系列剂量下给予的天然藻酸盐（分子量大于640,000）在小鼠和兔子中免疫原性较差，但该偶联物诱导产生了与天然藻酸盐结合的高水平抗体。兔子（而非小鼠）还产生了抗毒素免疫球蛋白抗体反应。源自其他三株铜绿假单胞菌的藻酸盐与同源的3064藻酸盐竞争结合抗偶联物抗体。这表明该偶联物引发了能够识别异源藻酸盐的抗体反应。用该偶联物免疫的兔子血清能有效促进人多形核白细胞对铜绿假单胞菌黏液样菌株的摄取和杀伤。相比之下，用天然藻酸盐免疫未产生调理抗体反应。兔抗偶联物抗体也中和了毒素A的细胞毒性潜力。
