Mouse Light Chain Research Articles

A Cytotoxic Antiglobulin Technique for Assay of Antibodies to Histocompatibility Antigens

Summary A method for increasing the cytotoxic activity of antibodies to lymphoid cells has been developed. Mouse isoantisera with 7 S and 18 S antibody activity were tested against an allogeneic lymphoid tumor cell. A variety of heterologous antiglobulins with different specificites were examined for their ability to produce inceased cytotoxicity. Both anti-IgG and anti-light chain reagents increased the cytotoxic titers of the 7 S antibody as much as 16-fold. Antibodies against mouse light chains were also able to enhance the cytotoxicity of the 18 S antibody. The degree of enhancement of cytotoxicity was considerably less than that found for antiglobulin reactions with erythrocytes and the possible explanations for these differences were discussed.

Genetic Control of Specific Immune Responses

This chapter discusses the genetic control of specific immune responses. The chapter reviews the recent important findings concerning “immune response genes” to antigenic determinants of different amino acids. The immune response to a specific antigen is a complex process that must involve genetic control at various levels. The fact that genetic factors are involved in the response to antigenic stimulus has long been known. The use of synthetic polypeptide antigens played a major role in elucidating the multiple genes that are described in the chapter. The intriguing question of at what level in the immune response these genes act remain to be determined. Genetic and structural analysis of normal immunoglobulins, myeloma proteins, and antibodies has produced a great deal of information about the genetic basis of antibody structure but has not yet given a clear picture of the genetic basis of antibody specificity. Structural analysis of myeloma proteins has led to a similar conclusion for the human and mouse light chain. The chapter summarizes the characteristic features common to the various genetic systems and analyzes the functions controlled by “immune response genes.”

The Specific Cleavage of Immunoglobulin Polypeptide Chains at Cysteinyl Residues

Abstract A method has been developed for fragmenting immunoglobulin light and heavy polypeptide chains at the cysteinyl residues that participate in intrachain disulfide bridges. The method involves the quantitative blockage of positive charges associated with lysyl and arginyl residues, the reduction and subsequent aminoethylation of disulfide bridges, and the treatment of aminoethylated, blocked chains with trypsin to yield the desired blocked peptide fragments. Lysine and arginine are modified by succinic anhydride and cyclohexanedione respectively, to yield stable trypsin-insensitive residues. The blocked peptides of heavy and light chains are separated on the basis of size by gel filtration chromatography in either 8 m urea or 0.01 m ammonium bicarbonate. The blocked peptides from a k-type Bence-Jones protein can be resolved into six chromatographic peaks. The peaks, in order of their elution, have been assigned loci within the known over-all structure of a Bence-Jones protein: E and D have not been characterized and probably represent partial digestion products; C and C' probably correspond to regions between the two intramolecular disulfide bridges; B contains the peptide corresponding to the switch region as judged by amino-terminal analysis and amino acid composition; A contains the carboxyl-terminal cysteinyl residue and corresponds to peptide fragments from the amino and carboxylterminal regions of the starting Bence-Jones protein. The peptide patterns obtained from whole normal mouse and human light chain are closely similar to the Bence-Jones profile and strongly suggest that the disulfide bridge pattern found for Bence-Jones proteins applies to the majority of κ and λ chains in the whole normal light chain population. A comparison of the gel filtration patterns of a γ2a mouse myeloma heavy chain and whole normal mouse heavy chains also indicates that a single heavy disulfide bridge pattern is largely conserved in the normal population. The similarity found between heavy and light chain peptide profiles supports the idea that heavy chains and light chains arose from a common ancestral gene as a result of gene duplications and mutations.

Light chains of rabbit immunoglobulin: assignment to the kappa class.

Normal rabbit gamma globulin was reduced under conditions presumed to break only interchain disulfide bridges, and the reduiced product was then blocked with C(14)-iodoacetamide. The light chains were separated from the heavy chains and subjected to peptic digestion. Two radioactive peptides were recovered from the digest. The peptides are apparently overlapping and represent the carboxyterminuis. Comparison of this region in the rabbit light chains with the corresponding amino acid sequences in various mouse and human light chains indicates that the rabbit light chains are of the (K)-class.

Location of Specificity and Allotypic Amino Acid Residues in Antibody Fd Fragments

Because of both the heterogeneity and limited supply of purified antibody, structural studies have been carried out on the homogeneous myeloma proteins which may belong to any one of the major classes of immunoglobulins but which lack antibody activity. Comparative analyses of human myeloma light chains by Putnam and Easley (1965), Hilschmann and Craig (1965), Titani, Whitley, Avogardo, and Putnam (1965), and Milstein (1966), and of mouse light chains by Hood, Gray, and Dreyer (1966) have provided the dramatic result of an amino-terminal half of variable sequence and a carboxy-terminal half of nearly invariant sequence. A similar structure appears likely for the heavy chain from the amino-terminal analyses of Wilkinson, Press, and Porter (1966) and the sequence determination of the common carboxyterminal portion by Hill, Delaney, Lebovitz, and Fellows (1966).