Naim J O, van Oss C J
Department of Surgery, Rochester General Hospital, New York 14621.
Immunol Invest. 1991 Jul;20(4):351-64. doi: 10.3109/08820139109057761.
Tuftsin (Thr-Lys-Pro-Arg) is a naturally occurring tetrapeptide that stimulates most known functions of the polymorphonuclear leukocyte and macrophage cell lines. We previously reported our unsuccessful attempts to generate antituftsin antibodies by conjugating tuftsin to several carrier proteins and by polymerizing the peptide with glutaraldehyde. To render tuftsin antigenic the following modifications were made to native tuftsin: three glycine residues were added to the N terminus of tuftsin (Gly3-tuf) and cysteine was added to the N terminus (Cys-tuf) and to the C terminus (tuf-Cys). Native tuftsin was covalently conjugated to sheep red blood cells (SRBC). In a separate experiment Balb/c mice primed with SRBC were immunized with 10(7) SRBC peptide conjugate. Native tuftsin and Gly3-tuf were also conjugated to keyhole limpet hemocyanin (KLH). In another experiment KLH and cationized bovine serum albumin (cBSA) were activated with sulfo-succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate (s-SMCC), which was used to control orientation of tuf-Cys and Cys-tuf when conjugated to each carrier protein. All conjugates were administered in complete Freund's adjuvant (CFA) except for cBSA conjugates which were administered in alum. Antibody response was determined by solid phase radioimmunoassay. Results showed that specific antituftsin antibodies were elicited only by Cys-tuf, conjugated to KLH. This study reaffirms that tuftsin is weakly antigenic and confirms the previous work by Gottlieb et al. that antibody to tuftsin can only be elicited when tuftsin is conjugated to the carrier protein KLH in a manner that leaves the peptide carboxyl end free.
促吞噬素（苏氨酸-赖氨酸-脯氨酸-精氨酸）是一种天然存在的四肽，可刺激多形核白细胞和巨噬细胞系的大多数已知功能。我们之前报道了通过将促吞噬素与几种载体蛋白偶联以及用戊二醛使该肽聚合来生成抗促吞噬素抗体的尝试未成功。为使促吞噬素具有抗原性，对天然促吞噬素进行了以下修饰：在促吞噬素的N末端添加了三个甘氨酸残基（Gly3-促吞噬素），并在N末端添加了半胱氨酸（Cys-促吞噬素）以及在C末端添加了半胱氨酸（促吞噬素-Cys）。天然促吞噬素与绵羊红细胞（SRBC）共价偶联。在另一项实验中，用SRBC致敏的Balb/c小鼠用10⁷个SRBC肽缀合物进行免疫。天然促吞噬素和Gly3-促吞噬素也与钥孔血蓝蛋白（KLH）偶联。在另一项实验中，用磺基琥珀酰亚胺4-(N-马来酰亚胺甲基)环己烷-1-羧酸盐（s-SMCC）激活KLH和阳离子化牛血清白蛋白（cBSA），当与每种载体蛋白偶联时，该试剂用于控制促吞噬素-Cys和Cys-促吞噬素的取向。除了在明矾中给药的cBSA缀合物外，所有缀合物均在完全弗氏佐剂（CFA）中给药。通过固相放射免疫测定法测定抗体反应。结果表明，仅通过与KLH偶联的Cys-促吞噬素引发了特异性抗促吞噬素抗体。这项研究再次证实促吞噬素的抗原性较弱，并证实了戈特利布等人之前的工作，即只有当促吞噬素以使肽羧基末端游离的方式与载体蛋白KLH偶联时，才能引发抗促吞噬素抗体。
