Iron oxide particles of average size 0.5-1.5 microns, covered by a silane coat carrying amino groups (Bio-Mag, Advanced Magnetics, Boston), were derivatized by reaction with N-[(gamma-maleimidobutyryl)oxy]-succinimide (GMBS), N-hydroxysuccinimidyl iodoacetate (NHIA), 2-iminothiolane (2-It), or N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP). The derivatized particles were suitable for the reaction with sulfhydryl groups and subsequently coated with monoclonal antibodies (MoAbs) of different classes and isotypes (IgM, IgG1, IgG2a, IgG2b, IgG3) as well as polyclonal rabbit anti-mouse IgG (RAM). The antibodies were reduced by dithiothreitol (DTT) and covalently conjugated to the BioMag derivatives via liberated sulfhydryls of the hinge region. The observed conjugation ratios, expressed as protein/iron (micrograms/mg), could be reproducibly varied for optimization. These ratios were dependent on the type and amount of antibody offered for coupling to the derivatized particles, decreasing as follows: polyclonal = IgM greater than IgG2b greater than IgG2a = IgG3 greater IgG1. The conjugation ratios were also dependent on the type and amount of the spacer used to derivatize the BioMag particles, decreasing as follows: GMBS greater than NHIA greater than 2-It greater than SPDP. The magnetically responsive magnetite-antibody conjugates ("magneto-beads"), carrying MoAb BMA 081 (anti-CD8; IgG2a), MoAb BB10 (anti-CD10/CALLA; IgG2b), MoAb VIL-A1 (anti-CD10; IgM), and polyclonal RAM, coupled similarly via 3.6 mumol of GMBS spacer per mg of Fe, were further investigated with respect to a depletion effect on specific cell subsets. The rates of cell depletion were found to be strongly dependent on the individual characteristics of the antibody used.(ABSTRACT TRUNCATED AT 250 WORDS)
A patient with lambda Bence-Jones proteinuria, Waldenström's macroglobulinaemia, and Franklin's disease (gamma HCD), but without clinical evidence of a lymphoproliferative disorder, is presented. The serum contained two distinct immunoglobulin abnormalities: a monoclonal immunoglobulin M (IgM) of lambda type, and a protein fragment which was immunologically related to immunoglobulin G (IgG) and devoid of light chain activity. This gamma HCD protein belongs to the gamma 3 subclass with a molecular weight of approximately 60,000 daltons. The urine contained a Bence-Jones lambda protein as well as the gamma HCD fragment. The two paraproteins were probably secreted by two different malignant clones. Ultrastructural study revealed pathological vacuolated plasma cells of a sort that has hitherto been principally described in association with micron HCD. The mechanism of the intracellular storage of pathological immunoglobulins is discussed in the light of the ultrastructural study.
Lactoferrin has been proposed recently as a physiological regulator of the granulocyte-monocyte progenitor (CFU-GM). This glycoprotein, when saturated with iron, has been said to limit the CFU-GM growth by decreasing production and release of colony stimulating activity by monocytes and macrophages. Human milk lactoferrin saturated with iron, at concentrations ranging from 10(-8) M, was added either to endogenously stimulated bone marrow cells or to mononucleated cells used as feeder layers for adherent cell-depleted marrow. Irrespective of the concentration of lactoferrin within the culture system used, no significant inhibition of the CFU-GM growth was observed. Moreover, the CFU-GM stimulating activity of medium conditioned by a 4 day incubation of 1 X 10(6) mononucleated blood cells in the presence or in the absence of lactoferrin was the same. Various possible explanations for not confirming the reported inhibiting activity of iron-saturated lactoferrin were explored: (a) masking inhibition of the system by prostaglandin E2 (PGE2), (b) masking inhibition of the system by bovine lactoferrin present in the fetal calf serum, (c) preinhibition of the system by leukemic-associated inhibitory activity possibly present in the culture system, (d) the iron and calcium content of the culture medium used, (e) the fixation of lactoferrin to plastic compounds, (f) the source of the human lactoferrin used, and (g) the marrow cell separation methods used. None of these factors was shown to play a role in vitro in the activity of lactoferrin and thus no evidence was found for a significant role of lactoferrin in the regulation of human granulopoiesis.
We compared the usefulness of four serum assays for classifying patients originally suspected of having an acute myocardial infarction. One of these is the long-used measurement of total creatine kinase (CK) activity. The other three are relatively new immunoassays: myoglobin by RIA, CK-BB by RIA, and CK-MB by immunoinhibition. When we evaluated test effectiveness with use of conventionally derived reference ranges, the results were misleading. However, by using receiver operating characteristic curves, we were able to effectively compare the four tests at all possible decision levels, rather than at only one. Multiple closely sequential serum specimens were obtained during the first four days after the onset of chest pain. Total CK, CK-MB, and CK-BB all behaved similarly, reaching peak diagnostic effectiveness at 18-20 h, when all three correctly classified 95% of the infarct patients, with a zero false-positive rate. However, total CK was more useful in identifying infarcts later in their courses than were the two CK isoenzyme tests. Myoglobin assay was most effective earlier in the course, at about 7 to 8 h. Our results indicate (a) that the tests for myoglobin and for CK or its isoenzymes are complementary and (b) that of the three CK tests, measurement of total CK activity provides the most information over the broadest segment of a patient's course.
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