Immunohistochemical analysis of human peripheral blood and lymphoid tissues using monoclonal antibodies immunoreactive with non-lymphoid cells

Summary Immunoperoxidase methods were used to study human peripheral blood and lymphoid tissues using a panel of monoclonal antibodies to non-lymphoid cells. The majority of peripheral blood monocytes were immunoreactive for LeuM1, LeuM2, LeuM3 and LeuM5. Rare peripheral blood monocytes were immunoreactive for R4/23. The different antibodies showed characteristic patterns of immunoreactivity in peripheral lymphoid tissues. LeuM1 was immunoreactive with scattered cells in the paracortex of lymph node and tonsil and with many cells in the marginal zone of the spleen. LeuM2 was immunoreactive with endothelial cells in lymph node and tonsil. A few cells in the red pulp of the spleen were immunoreactive for LeuM2. LeuM3 and R4/23 showed distinctive immunoreactivity in germinal centers of secondary follicles, giving a lacy pattern. LeuM3 was also immunoreactive with endothelium in lymph node and tonsil and with sinus lining cells in lymph node. LeuM5 was immunoreactive with macropages in the germinal center, fibroblastic reticulum cells in the mantle zone and interdigitating reticulum cells in the paracortex of lymph node and tonsil.     

人体胸腺和周围淋巴器官内T细胞亚群和NK细胞分布的研究

本文用多种Ｔ细胞和ＮＫ细胞单抗和免疫组织化学的ＡＢＣ技术,在冰冻切片上对人扁桃体、淋巴结、牌和胸腺内Ｔ细胞亚群和ＮＫ细胞的分布进行了检测。结果显示,ＣＤ５、ＣＤ８、ＣＤ４、ＣＤ３和ＡＩＧ３阳性细胞主要分布在扁桃体,淋巴结的副皮质区、脾的动脉周围淋巴鞘和胸腺,但各种抗体的反应强度不同。从各种Ｔ细胞工群的染色强度和形状看,胸腺髓质部的胸腺细胞相当于周围淋巴器官内的胸腺依赖区。胸腺内Ｔ细胞在分化过程中,质膜上的抗原也有相应变化。ＮＫ细胞主要分布在淋巴小结的生发中心,淋巴结和扁桃体的副皮质区,脾的红髓以及胸腺的筋质部。这些不同的分布,说明ＮＫ细胞不仅与淋巴小结的活动有关,可能还参与机体的免疫调节功能。     

Analysis of lymphoid and dendritic cells in human lymph node, tonsil and spleen. A study using monoclonal and heterologous antibodies

The distribution of lymphoid and dendritic cells in human reactive lymph nodes, tonsils and spleens was examined by means of an indirect immunoperoxidase technique, using a panel of monoclonal and heterologous antibodies. The antibodies used were directed against antigens present on T cell subsets (Leu1, leu2a, Leu3a, TA1, OKT6), various types of B cells (BA1, BA2, HLA-DR, CR1) and cells of the mononuclear phagocyte system (alpha HM1, TA1, CR1, OKM1, NA 1/34). In the lymph node and tonsil Leu3a-positive cells (T-helper/inducer phenotype) and Leu2a-positive cells (T-suppressor/cytotoxic phenotype) are found in the thymus-dependent or T-cell area; in the spleen Leu3a-positive cells are found mostly in the periarteriolar lymphocyte sheath (PALS), while Leu2a-positive T-suppressor/cytotoxic cells are almost completely restricted to the cords of Billroth in the red pulp. The cells in the mantle zone of germinal centres and in the primary follicles in lymph nodes, tonsils and spleens have B-cell properties (BA1-, HLA-DR-, and CR1-positive). The cells in the germinal centres show a similar staining pattern (HLA-DR-, and partly CR1-positive). Follicles and T-cell-dependent areas have specific dendritic cells, each with a specific staining pattern: the dendritic reticulum cell (DRC) of the follicle stain with CR1, HLA-DR, BA2 and alpha HM1; the interdigitating cell of the T-cell areas in the lymph node, tonsil and spleen stain with HLA-DR and BA1. Moreover, large dendritic OKT6-positive cells are found in the T-cell areas of some of the peripheral lymph nodes, and are probably Langerhans cells. It is concluded that human lymph nodes and tonsils have an identical compartimentalisation, clearly differing from the spleen in cellular organization.     

Immunoreactivity for S-100 protein in dendritic and in lymphocyte-like cells in human lymphoid tissues

S-100 protein is an immunohistochemical marker for a subset of dendritic cells, the interdigitating reticulum cells (IDRCs), which are mainly located in T-dependent areas of lymphoid tissues. In the present study we have investigated the distribution of S-100-positive cells in lymph nodes, spleen, thymus and peripheral blood of normal subjects. Immunoreactivity for S-100 protein was demonstrated in large cells with dendritic morphology and in small lymphocyte-like cells present in the lymph node paracortex, thymic medulla, splenic periarterial lymphatic sheaths (PALS) and in peripheral blood. S-100-positive lymphocyte-like cells were frequently detected around high endothelial venules (HEV) and were present in numbers comparable to those of S-100-positive IDRCs. Immunoelectron microscopy confirmed the existence of positive cells with lymphoid morphology and revealed that the intracellular distribution of the immunoreaction product was similar in lymphoid and dendritic cells. Further characterization of S-100-positive cells demonstrated that both lymphoid and dendritic cells were unreactive with a large panel of monocytic and macrophage markers.     

Morphology of germ-free piglets

The postnatal ontogeny of primary (thymus) and secondary (spleen, lymph node, lingual tonsil) lymphoid tissues was studied in germ-free colostrum-deprived piglets up to age of 68 days. The thymus, which is morphologically fully developed by the end of gestation, showed no significant differences in the germ-free and conventional state. In germ-free piglets, slow development of periarteriolarly organized lymph follicles occurred in the spleen up to the end of the observation period. As distinct from the conditions in the spleen of conventional animals, the presence of a large number of pyroninophilic cells was not observed in germ-free piglets and no germinal centres were found. A similar situation was seen in the mesenteric lymph nodes, in which, in conventional piglets, cells belonging to the plasmacyte series, as well as the germinal centres, proliferate by the 13th day. Differences were also found in the organization of the follicular lymphoid tissue in the wall of the terminal ileum. In germ-free piglets, the lymph follicles increased only very slowly in size during the observation period and germinal centres were absent, while in conventional piglots germinal centres were present from the 12th day. The view is expressed that the intestinal lymphoid tissue ought rather to be classified as peripheral lymphoid tissue.     

Occurrence of cysteine proteinase inhibitors in cells of the monocytic-histiocytic system. An immunohistochemical investigation

Human tissues are known to contain two low molecular weight (MW about 12,000) cysteine proteinase inhibitors, i.e. an acid inhibitor (ACPI) with pI 4.7-5.0 and a neutral inhibitor (NCPI) with pI 6.0-6.5. ACPI is abundant in cornifying epithelial tissues and in the dendritic reticulum cells of germinal centres of the lymph nodes. NCPI is abundant in lymphatic tissue and is known to be synthesized and released by mononuclear phagocytes. In this report NCPI was localized immunohistochemically in the epitheloid cells of most sarcoidotic lymph nodes, in lymph node macrophages after lymphangiography and in alveolar macrophages, while no ACPI could be demonstrated in the same cells by similar methods. These inhibitors were not demonstrable in lymph node sinus histiocytosis. Peripheral blood monocytes did not exhibit any NCPI immunoreactivity. In occasional blood monocytes anti-ACPI serum gave a weak reaction, the specificity of which is questionable. These data suggest that studies on cysteine proteinase inhibitors reveal basic differences in the various histiomonocytic cells and possibly differences in their functional stages.     

Cellular distribution of prothymosin alpha and parathymosin in rat thymus and spleen

By means of immunohistochemical methods, we have investigated the cellular distribution of prothymosin alpha and parathymosin in rat thymus and spleen, using specific antibodies raised against thymosin alpha-1 and against parathymosin. We observed prothymosin alpha immunoreactivity in lymphoid cells both in thymus and spleen. In the thymus, prothymosin alpha staining was more marked in cortex than in medulla. In the spleen, prothymosin alpha was found in lymphocytes of the periarteriolar lymphatic sheaths and was especially prominent in the germinal centers. Parathymosin immunoreactivity in the thymus was mainly localized in the medulla; positive cells were reticuloepithelial cells from the thymic reticulum and the blood barrier. Thymocytes were negative. In spleen, parathymosin was found in reticular cells arranged in a ring between the periarteriolar lymphatic sheath and the marginal zone. Our results do not support an exclusive role for these peptides as immune system hormones or cytokines.     

Distribution of cells bearing the Tac antigen during ontogeny of human lymphoid tissue

The monoclonal antibody anti-Tac, which binds to the interleukin 2 (IL 2) receptor, was used to identify this antigen in human fetal and adult lymphoid tissue. Liver, spleen, thymus, lymph node, and peripheral blood were examined for Tac-positive cells with the use of frozen sections or cytocentrifuge preparations. The results show that cells in the fetal and neonatal thymus express the Tac antigen; these cells are predominantly located in the medulla. The liver and spleen of both fetus and adult exhibit very few Tac-positive cells. Double staining demonstrates that cells bearing the Tac-antigen stain with Leu-4, an anti-T cell antibody. In adult lymph node tissue, the Tac-bearing cells are predominantly distributed in the interfollicular area, with positive cells also present in the germinal center and mantle zone. The Tac antigen is present on both T and B cells. Few Tac-positive cells are present in the circulating peripheral blood.     

Acute SIV Infection in Sooty Mangabey Monkeys Is Characterized by Rapid Virus Clearance from Lymph Nodes and Absence of Productive Infection in Germinal Centers

Lymphoid tissue immunopathology is a characteristic feature of chronic HIV/SIV infection in AIDS-susceptible species, but is absent in SIV-infected natural hosts. To investigate factors contributing to this difference, we compared germinal center development and SIV RNA distribution in peripheral lymph nodes during primary SIV infection of the natural host sooty mangabey and the non-natural host pig-tailed macaque. Although SIV-infected cells were detected in the lymph node of both species at two weeks post infection, they were confined to the lymph node paracortex in immune-competent mangabeys but were seen in both the paracortex and the germinal center of SIV-infected macaques. By six weeks post infection, SIV-infected cells were no longer detected in the lymph node of sooty mangabeys. The difference in localization and rate of disappearance of SIV-infected cells between the two species was associated with trapping of cell-free virus on follicular dendritic cells and higher numbers of germinal center CD4⁺ T lymphocytes in macaques post SIV infection. Our data suggests that fundamental differences in the germinal center microenvironment prevent productive SIV infection within the lymph node germinal centers of natural hosts contributing to sustained immune competency.     

Rat T lymphocyte-specific antigens and their cross-reactivity with mouse T cells.

Anti-rat T lymphocyte serum (ATLS)2 was prepared by immunizing rabbits with purified T cells from rat mesenteric nodes and absorbed with rat red cells and syngeneic sarcoma cells. The specificity of ATLS for rat T cells was confirmed by the following reasons: a) ATLS was not toxic for bone marrow cells but lysed most of the thymocytes and a number of spleen and lymph node cells, which were inversely correlated to the percentage of cells with B cell characteristics in respective organs; b) anatomical localization of ATLS-reactive cells in lymphoid organs coincided to the thymus-dependent areas, i.e. the paracortex of lymph node and the periarteriolar region of spleen; c) spleen cells treated with ATLS and complement failed to respond to phytohemagglutinin but normally responded to bacterial lipopolysaccharide; d) those cells treated with ATLS and complement could not induce a graft-vs-host reaction in F1 hosts, whereas the same treatment did not affect direct plaque-forming cells. All of these data confirm the specificity of ATLS and indicate that ATLS recognizes rat T lymphocyte-specific antigens (RTLA). Absorption studies showed that RTLA were present in higher concentration on medullary thymocytes and peripheral T cells than on cortical thymocytes, but absent from bone marrow, liver, and brain tissues. When the cross-reactivity of RTLA with mouse T cells was studied by C-dependent cytotoxicity and immunofluorescence, it was found that mouse T cells shared at least one determinant of RTLA with rat T cells, and that distribution pattern of the cross-reacting antigens in mouse lymphoid tissues was essentially the same as that of RTLA in rat lymphoid organs.     

B cell-specific S1PR1 deficiency blocks prion dissemination between secondary lymphoid organs

Many prion diseases are peripherally acquired (e.g., orally or via lesions to skin or mucous membranes). After peripheral exposure, prions replicate first upon follicular dendritic cells (FDC) in the draining lymphoid tissue before infecting the brain. However, after replication upon FDC within the draining lymphoid tissue, prions are subsequently propagated to most nondraining secondary lymphoid organs (SLO), including the spleen, by a previously underdetermined mechanism. The germinal centers in which FDC are situated produce a population of B cells that can recirculate between SLO. Therefore, we reasoned that B cells were ideal candidates by which prion dissemination between SLO may occur. Sphingosine 1-phosphate receptor (S1PR)1 stimulation controls the egress of T and B cells from SLO. S1PR1 signaling blockade sequesters lymphocytes within SLO, resulting in lymphopenia in the blood and lymph. We show that, in mice treated with the S1PR modulator FTY720 or with S1PR1 deficiency restricted to B cells, the dissemination of prions from the draining lymph node to nondraining SLO is blocked. These data suggest that B cells interacting with and acquiring surface proteins from FDC and recirculating between SLO via the blood and lymph mediate the initial propagation of prions from the draining lymphoid tissue to peripheral tissues.     

Three distinct antigen systems on human B cell subpopulations as defined by monoclonal antibodies

Three novel antigen systems (L22, L23, and L24) expressed on human B cell subpopulations were identified by using TB1-2C3, TB1-2B3, and TB1-3C1 monoclonal antibodies, respectively. L22 was expressed on a minor subpopulation of B cells in human lymphoid tissues and in the peripheral blood. These B cells associated with L22 were resting small B cells mainly located in the mantle zone of lymphoid follicles, most of which also expressed IgM and IgD on their cell membrane. This antigen was absent from all cultured hemopoietic cell lines including B cell-derived cell lines as well as from all human B cell malignancies, except for B cell-type chronic lymphocytic leukemia and hairy cell leukemia. L23 and L24, on the other hand, existed on approximately two-third of B cells in blood and lymphoid tissues. These L23 and L24 antigens were expressed largely on small lymphocytes located in the mantle zone of lymphoid follicles and to a lesser extent on large blastic cells within lymphoid germinal centers. L23 and L24, like L22, seem to disappear from B cells during their differentiation into antibody-secreting cells, because they were not expressed on normal and neo-plastic plasma cells. This is additionally confirmed by the observation that L23 and L24 were expressed little or not at all on pokeweed mitogen-activated and Epstein-Barr virus-transformed B cells, and were absent from some of B cell malignancies that have been thought to correspond to the later stages of B cell development. Although L23, but not L22 and L24, was faintly expressed on mature granulocytes and monocytes, none of L22, L23, or L24 existed on human thymus and T cells. Immunoprecipitation studies showed that L23 and L24 were different molecular species consisting of a single glycoprotein with m.w. of 205,000 and 145,000, respectively. L22 antigen is presently under study.     

A study of lymphocyte subsets in human normal tonsil and lymph node

A series of T and B lymphocyte specific monoclonal antibodies was used to determine the localization of lymphocyte subpopulation in frozen and paraffin tissue sections of human normal tonsil and lymph node by means of immunocytochemical technique. In the paracortical and interfollicular area of tonsil and lymph node, most lymphocytes reacted with Leu 1, Leu 3 a, Leu 4 and OKT4. The numbers of Leu 2 a and OKT8 positive cells were rare in tissue. These cells were not only limited in paracortical area, they also appeared in considerable numbers in medullary cords of lymph nodes. Leu 2 a and OKT 8 positive cells decreased with prominent follicular hyperplasia of tonsils. In addition, substantial leu 3 a and Leu 4 cells were found in the germinal centers. This finding supports the importance of these lymphocyte subsets in regulation of human immune response. In the mantle zone of secondary follicles, the majority of lymphocytes were positive for OKB 2 and BA 1, whereas, the IgM positive cells were predominately observed in the cytoplasma and extracellular substance of B lymphocytes in the germinal centers, but the lymphocytes bearing sIgM were rarely observed. In the mantle zone, the IgM were frequently found on the surface of membrane of small lymphocytes, however, the staining intensity was much than that in the germinal centers.     

Human ferritin: effects of antigen source and fixation on leucocyte staining by immunoperoxidase technique

The localization of ferritin was studied in peripheral blood cells and variously fixed tissues with the antibodies against ferritins isolated from human heart and spleen. The unlabelled antibody enzyme method (PAP) was used to detect the binding sites of antibodies. In peripheral blood cell smears both antisera gave rise to strong staining of polymorphonuclear (PMN) cell cytoplasm, whereas the monocytes stained relatively weakly. There were no staining differences between the two antisera. In human spleen sections the spleen ferritin antiserum stained the PMN cells and sinusoidal lining cells, whereas the heart ferritin antiserum stained only PMN cells. Neither of the two antisera stained monocytes in the spleen sections. This finding was observed in specimens fixed in Bouin's fixative, Baker's fixative and neutral formalin. However, the immunoreactivity of ferritin was totally destroyed by some other fixatives (Carnoy's fixative, formol sucrose and glutaraldehyde). These results suggest that ferritin is more readily released from monocytes than from PMN cells, and that mature spleen macrophages contain antigenic determinants of ferritin that are recognized only by anti-spleen ferritin antiserum.     

Comparative migration of B- and T-Lymphocytes in the rat spleen and lymph nodes.

B-lymphocytes were obtained either by thoracic duct cannulation of thymectomized, irradiated rats or by isolation of complement-receptor-bearing lymphocytes from normal rats. They were labeled in vitro with [³H]-leucine and injected iv into syngeneic recipients from which samples of spleen and lymph node were taken at intervals from 15 min to 48 hr after injection. The sites of initial localisation of B- and T-lymphocytes were identical suggesting that the cells migrated into both organs by a common entrance. The two cell types remained closely associated for several hours in the paracortex of lymph nodes and at the periphery of the periarteriolar lymphoid sheath of the spleen. After 1–6 hr, B-cells segregated from T-cells by moving on into the adjacent part of the lymphocyte corona in the follicular area. By 24 hr, B-cells were evenly distributed throughout the corona. A definite minority of B-cells but no T-cells were seen within the germinal centres. In the spleen, T-cells moved into the central area of the periarteriolar sheath before returning to the blood. The immunological significance of the routes of B- and T-cell migration is discussed.     

In vitro development of a primary antibody response with lymph node cells.

Utilizing a variety of lymphoid tissues from three common laboratory species, comparative studies were performed to investigate the competence of the dissociated cells to respond to a heterologous erythrocyte with the development of specific plaque-forming cells. Dissociated spleen cells harvested from BDF1 mice consistently developed specific plaque-forming cells (PFC) to sheep red blood cells (SRBC), while hamster spleen cells inconsistently developed specific antibody-forming cells to SRBC. Under identical conditions, guinea pig spleen cells did not develop significant numbers of PFC to SRBC. However, lymph node cell cultures of all three species tested yielded specific PFC. In the mouse and hamster lymph node cell cultures, the yield of PFC per culture or per 10⁶ recovered viable cells was always greater than the yield from companion spleen cell cultures. Guinea pig mesenteric lymph node cell cultures developed the major PFC response to SRBC, while both mesenteric and peripheral lymph node cell cultures from hamsters were equivalent in their response to SRBC. The data demonstrate that it is possible to develop a primary antibody response to SRBC in vitro utilizing normal endogenous hamster or guinea pig lymphoid cells, if lymph nodes are the source of cells.     

Antigenic relationship between bone marrow lymphocytes cortical thymocytes and a subpopulation of peripheral T cells in the rat: description of a bone marrow lymphocyte antigen.

Populations of rat bone marrow lymphocytes (BML) consisting of approximately 90 percent, “tnull” cells were prepared by density gradient centrifugation, passage through a column of fine glass beads, and treatment with anti-T cell and anti-B cell serum plus complement. Antisera to these bone marrow lymphocytes were raised in rabbits. After absorption with RBC and peritoneal exudate cells, the anti-BML sera were found by immunofluorescence to react selectively with “null” cells in bone marrow, with cortical thymocytes, and with a cortisone-sensitive subset of T cells in blood and in spleen, possibly in red pulp. The antigen that is common to these cell types is designated the rat bone marrow lymphocyte antigen (RBMLA). Lymphocytes that are positive fur KBMLA are negative for another lymphocyte-specific heteroantigen, rat musked thymocyte antigen (RMTA). As shown previously, RMTA is present on medullary thymocytes and ou cortisone-resistant T cells in white pulp of spleen, paracortex of lymph node and thoracic duct lymph. It is postulated that two developmentally and functionally distinct lines of T cells exist in peripheral lymphoid tissues of the rat, one derived from cortical thymocytes and one derived from medullary thymocytes. It is further postulated that the “null” population of bone marrow lymphocytes contains the lymphopoietic stem cells from which these two lines of T cells originate.     

Scanning, transmission and immunoelectron microscopical studies of the tonsil-like lymphoid organ of normal and horseradish-peroxidase-injected laboratory suncuses

Fine structures of normal and horseradish peroxidase (HRP)-stimulated tonsil-like lymphoid organs of the laboratory suncus (Suncus murinus) were examined by scanning, transmission and immunoelectron microscopies. The normal organs appear as a pair of small oval protrusions at the upper lateral sites of the fauces, and consist of a single lymph nodule with a germinal center and a crypt-like epithelium with prominent lymphoid cell infiltration. Postcapillary venules (PCV) and efferent lymphatics are associated within the marginal region of the nodule and separated from the neighboring pharyngeal tissues. Numerous lympho-plasma cells, interdigitating cells and reticulum cells occurred within the lymphoid parenchyma, as well as in the intraepithelial infiltrating cell populations. In the HRP-stimulated animals, the anti-HRP antibodies producing lympho-plasma cells were often seen in the parenchyma and epithelia; however, similar HRP-antibody-positive lymphocytes were rarely detected in the PCV lumina. In addition, some HRP antibody bearing interdigitating cells were also identified in the same parenchyma. These data indicate that the suncus' tonsil-like organs have a positive immune function to oral antigens, together with the suncus' systemic immune system and it is hypothetically presumed that the organ may correspond to a homologous organ of the human palatine tonsil in comparative anatomy.     

Immunohistological localization of alpha 1-microglobulin in normal rat tissues

The tissue distribution of rat alpha 1-microglobulin (alpha 1-m) was studied by indirect immunofluorescence in various rat tissues using a polyvalent rabbit antiserum to the purified antigen and a monoclonal antibody (H23) to the human homologue, in parallel with a polyclonal anti-rat IgA antiserum. It was found that all tissues stained by anti-IgA were also alpha 1-m positive; these tissues included tissues of the stomach, duodenum, ileum, colon, pancreas, trachea, esophagus and jejunum. However, the observation that IgA plasma cells as well as secretory cells, while positively stained by anti-IgA, are alpha 1-m negative suggests that the association between IgA and alpha 1-m occurs at a postsecretory stage, after the IgA molecules have been transported across the epithelial cells. Additionally, hepatocytes were intensely stained by anti-alpha 1-m antibodies, indicating that the liver, as already suggested by metabolic studies on isolated guinea-pig liver explants, may be responsible for the synthesis of this protein. Among lymphoid tissues, an intense and homogeneous staining was observed in the thymus and the white pulp of the spleen. Sections of lymph nodes, however, showed differential staining; apart from a few isolated dendritic cells in the mantle region of the lymphoid follicles, the germinal centers and medullary cords showed no staining with anti-alpha 1-m antibodies. The paracortical cells, macrophages in the subcapsular sinus, and interfollicular lymphocytes showed intense cytoplasmic staining with anti-alpha 1-m antibodies. In other tissues, macrophages, monocytes, tissue histiocytes, and dendritic cells were alpha 1-m positive. Although they confirm the presence of alpha 1-m in the lymphoid tissues, as already reported in man, these results show that the protein is also present in hepatocytes and in exocrine fluids containing IgA. Since alpha 1-m, like secretory component, can bind to IgA to form stable complexes, these two heavily glycosylated proteins may have similar biologic properties.     

Homing of germinal-center cells into germinal centers of lymph node via afferent lymphatics

Summary Affinity of lymphoid cells for the microenvironment of germinal centers (GC), as detectable in transfer experiments by rapid homing in spleen GC from the blood, is a capacity expressed by only a subset of lymphoid cells, in particular by those constituting a GC. However, when introduced into the blood stream, these cells do not home into GC of lymph nodes and gut-associated lymphoid tissues. To investigate further this homing inability for high endothelial venule (HEV)-containing lymphoid tissues, GC cells isolated from donor rabbit appendix were labeled in vitro with ³H-leucine and injected into an afferent lymph vessel of recipient popliteal lymph nodes. Draining lymph nodes were removed 15 min to 24 h after cell administration and prepared for radioautography. For reference, the migration of cells isolated from Peyer's patches and thoracic duct lymph was also studied. By use of appendix GC cells, large numbers of labeled cells were found to migrate into GCs of the outer cortex centripetally, i.e., from the subcapsular sinus through the lymphocyte corona into the GC proper. The same was observed for cells from Peyer's patches, although in smaller numbers. Thoracic duct lymphocytes were only localized in the lymphocyte corona and the deep cortex. Thus, appendix GC cells and a subpopulation of cells from Peyer's patches can reach lymph node GC, but only when administered intralymphatically. We conclude that cells expressing affinity for the GC microenvironment do so for both spleen and lymph node GC, but do not have the capacity to interact with the wall of HEV; its implication for the understanding of the dynamics of a GC reaction is discussed.Abbreviations GC germinal center - GCC germinal-center cells - AGCC appendix germinal-center cells - GCPC germinal-center precursor cells - GCSC germinal-center seeking cells - HEV high endothelial venules - SRBC sheep red blood cells - PP Peyer's patch - TDL thoracic duct lymphocytes - NCS newborn calf serum - PBS phosphate-buffered saline - PNA peanut agglutinin - LN lymph node - LC lymphocyte corona - DC deep cortex unit