Journal: Immunology and Cell Biology

Article Title: Genetic regulation of antibody responsiveness to immunization in substrains of BALB /c mice

doi: 10.1111/imcb.12199

Figure Lengend Snippet: Antibody responsiveness of BALB /c mice from different sources. (a) BALB /c A and BALB /c B mice were vaccinated subcutaneously with trivalent influenza vaccine and trivalent influenza vaccine‐specific IgG1 titers were measured 28 days after immunization by ELISA . BALB /c A and BALB /c B mice were vaccinated subcutaneously with NP‐OVA + IFA and (b) NP ‐specific IgG1, IgG2a and IgE and (c) total IgG1, IgG2a and IgE serum titers measured 14 days after immunization by ELISA . Data points represent individual mice and heights of the bar represent the median. Dashed lines represent lower limit of sensitivity, set at blank OD . * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001 and **** P ≤ 0.0001. Data are representative of at least two experiments.

Article Snippet: Serum NP‐specific antibody levels were determined with NP coupled to bovine serum albumin (NP‐BSA; Biosearch Technologies) for capture followed by biotinylated anti‐mouse IgG1, IgG2a, IgE or IgM detection.

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Immunology and Cell Biology

Article Title: Genetic regulation of antibody responsiveness to immunization in substrains of BALB /c mice

doi: 10.1111/imcb.12199

Figure Lengend Snippet: Microbiota composition does not regulate altered antibody responsiveness in BALB /c A and BALB /c B mice. BALB /c A and BALB /c B mice were cohoused to promote microbiota transfer and 4 weeks later vaccinated with NP ‐ OVA in IFA ; cohoused BALB /c A ( A CH ), BALB /c B ( B CH ), BALB /c A and BALB /c B. Gut microbiota composition was determined with taxonomic profiling of fecal bacterial communities from 16s rRNA sequencing. (a) Relative abundance at the phyla level with key populations‐of‐interest highlighted at the order, family or genus level and (b) principal component analysis of fecal microbiota composition; left and right plot provide two views of the taxonomic diversity. Samples taken 1 day prior to immunization ( n = 5 per group). NP ‐specific IgG1, IgG2a and IgE titers in serum of (c) A CH and BALB /c A and NP ‐specific IgG1, IgG2a and IgE titers in serum of (d) B CH and BALB /c B 14 days after immunization with NP ‐ OVA in IFA . Data points represent individual mice and heights of the bar represent the median. Dashed lines represent lower limit of sensitivity, set at blank OD . Data are representative of at least two experiments.

Article Snippet: Serum NP‐specific antibody levels were determined with NP coupled to bovine serum albumin (NP‐BSA; Biosearch Technologies) for capture followed by biotinylated anti‐mouse IgG1, IgG2a, IgE or IgM detection.

Techniques: Sequencing

Journal: Immunology and Cell Biology

Article Title: Genetic regulation of antibody responsiveness to immunization in substrains of BALB /c mice

doi: 10.1111/imcb.12199

Figure Lengend Snippet: Germinal center B cells from low‐responder BALB /c A mice possess a reduced class‐switch capability. BALB /c A and BALB /c B mice were vaccinated subcutaneously with NP‐OVA + IFA. Draining lymph nodes and serum were taken 14 days after immunization. (a) Formation of GC structures was visualized by immunofluorescence imaging; Cyan: GL 7, Green: CD 4, Magenta: B220. Representative images of one mouse per group. Scale bar 500 μm. (b) The number of GL7+ cell clusters within the B‐cell follicle counted per section. (c) NP ‐specific IgM and (d) IgG1 serum titers measured 14 days after immunization by ELISA . (e) Number of NP + GC B cells in the lymph node. (f) Frequency of NP + IgM+ GC B cells in the lymph node. (g) Dot plots illustrate IgG1 expression and NP ‐specificity within the GC B‐cell population; pre‐gated on size, viability CD 138– B220+ GL 7+ CD 38–. Representative dot plots of one mouse per group. (h) Frequency and (i) number of NP + IgG1+ GC B cells in the lymph node. (j) Number of NP + IgG1+ plasma cells in the lymph node; gated on size, viability CD 138+ B220–. (k) Number of IgG1+ plasma cells in the bone marrow 28 days after immunization with OVA adjuvanted with alum; gated on size, viability CD 138+ B220–. Data points represent individual mice and heights of the bar represent the median. Data are representative of at least three experiments.

Article Snippet: Serum NP‐specific antibody levels were determined with NP coupled to bovine serum albumin (NP‐BSA; Biosearch Technologies) for capture followed by biotinylated anti‐mouse IgG1, IgG2a, IgE or IgM detection.

Techniques: Immunofluorescence, Imaging, Enzyme-linked Immunosorbent Assay, Expressing, Clinical Proteomics

Journal: Immunology and Cell Biology

Article Title: Genetic regulation of antibody responsiveness to immunization in substrains of BALB /c mice

doi: 10.1111/imcb.12199

Figure Lengend Snippet: Isotype class switching is defective in BALB /c A B cells. Naïve splenic B cells of BALB/c A and BALB/c B mice were stimulated with LPS + IL ‐4 and monitored for (a) survival at 24 h and (b) C‐S to IgG1 and proliferation at 90 h by flow cytometry; graphs show the frequency and number of IgG1+ cells in each generation gated on size, viability B220+ CD 138+ CTV IgM– IgG1+. (c) Naïve splenic B cells were stimulated with α CD 40 + IL ‐4 and monitored for proliferation and C‐S to IgG1 at 90 h by flow cytometry; graphs show the frequency and number of IgG1+ cells in each generation gated on size, viability B220+ CD 138+ CTV IgM– IgG1+. (d) Naïve splenic B cells were stimulated with LPS and monitored for proliferation and C‐S to IgG3 at 90 h by flow cytometry; graphs show the frequency and number of IgG3+ cells in each generation gated on size, viability B220+ CD 138+ CTV IgM– IgG3+. (a) Each data point represents one sample. (b–d) Representative dot plots from one sample per group and data from triplicate cultures were graphed. Statistical significance was determined using an unpaired t ‐test. Data are representative of at least three experiments.

Article Snippet: Serum NP‐specific antibody levels were determined with NP coupled to bovine serum albumin (NP‐BSA; Biosearch Technologies) for capture followed by biotinylated anti‐mouse IgG1, IgG2a, IgE or IgM detection.

Techniques: Flow Cytometry