amca streptavidin  (Vector Laboratories)


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    Name:
    AMCA Streptavidin
    Description:
    AMCA Streptavidin can be used to detect biotinylated secondary antibodies and other macromolecules in applications such as immunofluorescence in situ hybridization or flow cytometry Vector Laboratories fluorochrome conjugated streptavidin and avidin reagents are highly purified and possess very low non specific binding properties They have extremely high affinity for biotin Amplification of fluorescent signals can be easily achieved with our biotinylated secondary antibodies followed by our highly purified fluorochrome labeled streptavidin or avidin Using a biotin avidin or biotin streptavidin detection system results in an additional layer of amplification over a directly conjugated secondary antibody AMCA Streptavidin is produced by conjugating streptavidin with a coumarin fluorescent dye 7 amino 4 methylcoumarin 3 acetic acid This derivative excites in the ultraviolet 350 nm and emits in the visible 450 nm producing an intense blue fluorescence Most paraffin embedded tissues have little autofluorescence under those conditions AMCA Streptavidin may provide an advantage in many labeling situations over conjugates of fluorescein and related dyes AMCA Streptavidin can be used on frozen or paraffin embedded sections as a single label or in combination with other fluorescent conjugates for multiple labeling studies
    Catalog Number:
    SA-5008
    Price:
    None
    Size:
    1 mg
    Category:
    Protein chemifluorescent detection reagents or kits or substrates
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    Structured Review

    Vector Laboratories amca streptavidin
    AMCA Streptavidin
    AMCA Streptavidin can be used to detect biotinylated secondary antibodies and other macromolecules in applications such as immunofluorescence in situ hybridization or flow cytometry Vector Laboratories fluorochrome conjugated streptavidin and avidin reagents are highly purified and possess very low non specific binding properties They have extremely high affinity for biotin Amplification of fluorescent signals can be easily achieved with our biotinylated secondary antibodies followed by our highly purified fluorochrome labeled streptavidin or avidin Using a biotin avidin or biotin streptavidin detection system results in an additional layer of amplification over a directly conjugated secondary antibody AMCA Streptavidin is produced by conjugating streptavidin with a coumarin fluorescent dye 7 amino 4 methylcoumarin 3 acetic acid This derivative excites in the ultraviolet 350 nm and emits in the visible 450 nm producing an intense blue fluorescence Most paraffin embedded tissues have little autofluorescence under those conditions AMCA Streptavidin may provide an advantage in many labeling situations over conjugates of fluorescein and related dyes AMCA Streptavidin can be used on frozen or paraffin embedded sections as a single label or in combination with other fluorescent conjugates for multiple labeling studies
    https://www.bioz.com/result/amca streptavidin/product/Vector Laboratories
    Average 94 stars, based on 182 article reviews
    Price from $9.99 to $1999.99
    amca streptavidin - by Bioz Stars, 2020-08
    94/100 stars

    Images

    1) Product Images from "A Barrel-Related Interneuron in Layer 4 of Rat Somatosensory Cortex with a High Intrabarrel Connectivity"

    Article Title: A Barrel-Related Interneuron in Layer 4 of Rat Somatosensory Cortex with a High Intrabarrel Connectivity

    Journal: Cerebral Cortex (New York, NY)

    doi: 10.1093/cercor/bht263

    Average axonal domains and firing patterns of the 3 different types of fast-spiking L4 interneurons. ( A 1–3) Representative Neurolucida reconstructions of L4 interneurons that were classified as cluster 1 ( A 1), cluster 2 ( A 2), and cluster 3 ( A 3) cells, respectively (axons are given in blue, the somatodendritic domain in red). ( B 1–3) Average axonal length density maps (blue) of the 3 different clusters, respectively. White, thin contour lines enclosing 70%, 80%, and 90% of the integrated axonal density are shown superimposed. The red triangles depict the location of the L4 interneuron somata with respect to the center of the barrel. Note that cluster 1 interneurons have axonal domains that project both throughout cortical layer 2/3, 4, and 5 with the highest density in layer 2/3. In contrast, cluster 3 interneurons have an axonal domain that resides almost exclusively within the home barrel. Cluster 2 interneurons constitute an intermediate class between cluster 1 and 3 neurons with an axon largely confined to the home barrel with only short vertically ascending axonal collaterals in lower layer 2/3. ( C 1–3) Examples of AP firing patterns elicited by rectangular current pulses injected at the soma of the 3 L4 interneuron types. The maximum firing frequency in all of these neurons was ∼300 Hz, that is, all 3 types can be classified as interneurons with FS characteristics. There were no significant differences in the firing characteristics and, thus, the L4 interneurons cannot be discriminated by these criteria. ( D ) Immunohistochemistry of a typical cluster 3 neuron. AMCA streptavidin was used to determine the location and morphology of the neuron, which was positive for PV but negative for CB.
    Figure Legend Snippet: Average axonal domains and firing patterns of the 3 different types of fast-spiking L4 interneurons. ( A 1–3) Representative Neurolucida reconstructions of L4 interneurons that were classified as cluster 1 ( A 1), cluster 2 ( A 2), and cluster 3 ( A 3) cells, respectively (axons are given in blue, the somatodendritic domain in red). ( B 1–3) Average axonal length density maps (blue) of the 3 different clusters, respectively. White, thin contour lines enclosing 70%, 80%, and 90% of the integrated axonal density are shown superimposed. The red triangles depict the location of the L4 interneuron somata with respect to the center of the barrel. Note that cluster 1 interneurons have axonal domains that project both throughout cortical layer 2/3, 4, and 5 with the highest density in layer 2/3. In contrast, cluster 3 interneurons have an axonal domain that resides almost exclusively within the home barrel. Cluster 2 interneurons constitute an intermediate class between cluster 1 and 3 neurons with an axon largely confined to the home barrel with only short vertically ascending axonal collaterals in lower layer 2/3. ( C 1–3) Examples of AP firing patterns elicited by rectangular current pulses injected at the soma of the 3 L4 interneuron types. The maximum firing frequency in all of these neurons was ∼300 Hz, that is, all 3 types can be classified as interneurons with FS characteristics. There were no significant differences in the firing characteristics and, thus, the L4 interneurons cannot be discriminated by these criteria. ( D ) Immunohistochemistry of a typical cluster 3 neuron. AMCA streptavidin was used to determine the location and morphology of the neuron, which was positive for PV but negative for CB.

    Techniques Used: Injection, Immunohistochemistry

    2) Product Images from "Netrin-1 promotes adipose tissue macrophage accumulation and insulin resistance in obesity"

    Article Title: Netrin-1 promotes adipose tissue macrophage accumulation and insulin resistance in obesity

    Journal: Nature medicine

    doi: 10.1038/nm.3467

    Netrin-1 promotes macrophage retention in adipose tissue during obesity ( a ) qRT-PCR analysis of mRNA for M1 ( Nos2, Tnfa , Il6 ) and M2 ( Cd206, IL10, Pparg ) markers in macrophages isolated from the VAT of mice fed chow ( n = 7) or HFD ( n = 9). (b) Analysis of macrophage recruitment and retention in VAT of mice fed a chow or HFD using a fluorescent bead-tracking model. Left: Representative images of fluorescent bead-labeled cells in VAT of HFD-fed mice at day 3 and 14. Scale bar = 100 µm. Right: Mean number of bead-labeled macrophages in VAT sections ( n = 5 mice per group). ( c ) F4/80 staining (red) of VAT from HFD-fed mice 3 and 14 days post-labeling showing fluorescent beads (green) in macrophages in crown-like structures. (d) Mean number of beads in mesenteric lymph nodes on days 3 and 14 ( n = 5 mice per group). Data in a, b, and d are the mean ± s.e.m. * P
    Figure Legend Snippet: Netrin-1 promotes macrophage retention in adipose tissue during obesity ( a ) qRT-PCR analysis of mRNA for M1 ( Nos2, Tnfa , Il6 ) and M2 ( Cd206, IL10, Pparg ) markers in macrophages isolated from the VAT of mice fed chow ( n = 7) or HFD ( n = 9). (b) Analysis of macrophage recruitment and retention in VAT of mice fed a chow or HFD using a fluorescent bead-tracking model. Left: Representative images of fluorescent bead-labeled cells in VAT of HFD-fed mice at day 3 and 14. Scale bar = 100 µm. Right: Mean number of bead-labeled macrophages in VAT sections ( n = 5 mice per group). ( c ) F4/80 staining (red) of VAT from HFD-fed mice 3 and 14 days post-labeling showing fluorescent beads (green) in macrophages in crown-like structures. (d) Mean number of beads in mesenteric lymph nodes on days 3 and 14 ( n = 5 mice per group). Data in a, b, and d are the mean ± s.e.m. * P

    Techniques Used: Quantitative RT-PCR, Isolation, Mouse Assay, Labeling, Staining

    Netrin-1 is expressed by adipose tissue macrophages in obese human VAT ( a ) qRT-PCR analysis of mRNA isolated from VAT of lean ( n = 5) or obese ( n = 5) human subjects. Data are the mean ± s.e.m. * P
    Figure Legend Snippet: Netrin-1 is expressed by adipose tissue macrophages in obese human VAT ( a ) qRT-PCR analysis of mRNA isolated from VAT of lean ( n = 5) or obese ( n = 5) human subjects. Data are the mean ± s.e.m. * P

    Techniques Used: Quantitative RT-PCR, Isolation

    Palmitate upregulates Ntn1 and Unc5b expression in macrophages ( a ) Ntn1 and Unc5b mRNA in BMDM treated with 250 µM BSA or palmitate for 24 h. ( b ) Ntn1 and ( c ) Unc5b promoter-luciferase reporter activity in HEK293T cells treated with 250 µM BSA or palmitate in the presence of the NFkB inhibitor BAY11-7082 (10 µmol/L) or vehicle (veh). ( d ) Concentration of netrin-1 in culture supernatants of BMDM treated with 250 µM BSA or palmitate for 24 h in the presence of BAY11-7082 or vehicle. ( e ) Ntn1 and Unc5b mRNA in BMDM treated with conditioned media (CM) from 3T3L1 adipocytes exposed to BSA or palmitate (250 µM) for 24 h. ( f ) qRT-PCR of mRNA from 3T3L1 adipocytes treated with BSA or palmitate (250µM) for 24 h. ( g ) Ntn 1 mRNA in BMDM treated with TNFα (10 ng/ml), IL-6 (40 ng/ml) or IL-4 (10 ng/ml) for 24 h. ( h ) Ntn1 and Unc5b mRNA in BMDM treated with CM from 3T3L1 adipocytes exposed to BSA or palmitate (250 µM) in the presence or absence of anti-TNFα, anti-IL-6 or both. Data presented are the mean ± s.d. of triplicate samples from a single experiment and are representative of three independent experiments. * P
    Figure Legend Snippet: Palmitate upregulates Ntn1 and Unc5b expression in macrophages ( a ) Ntn1 and Unc5b mRNA in BMDM treated with 250 µM BSA or palmitate for 24 h. ( b ) Ntn1 and ( c ) Unc5b promoter-luciferase reporter activity in HEK293T cells treated with 250 µM BSA or palmitate in the presence of the NFkB inhibitor BAY11-7082 (10 µmol/L) or vehicle (veh). ( d ) Concentration of netrin-1 in culture supernatants of BMDM treated with 250 µM BSA or palmitate for 24 h in the presence of BAY11-7082 or vehicle. ( e ) Ntn1 and Unc5b mRNA in BMDM treated with conditioned media (CM) from 3T3L1 adipocytes exposed to BSA or palmitate (250 µM) for 24 h. ( f ) qRT-PCR of mRNA from 3T3L1 adipocytes treated with BSA or palmitate (250µM) for 24 h. ( g ) Ntn 1 mRNA in BMDM treated with TNFα (10 ng/ml), IL-6 (40 ng/ml) or IL-4 (10 ng/ml) for 24 h. ( h ) Ntn1 and Unc5b mRNA in BMDM treated with CM from 3T3L1 adipocytes exposed to BSA or palmitate (250 µM) in the presence or absence of anti-TNFα, anti-IL-6 or both. Data presented are the mean ± s.d. of triplicate samples from a single experiment and are representative of three independent experiments. * P

    Techniques Used: Expressing, Luciferase, Activity Assay, Concentration Assay, Quantitative RT-PCR

    Netrin-1 blocks chemokine-induced migration of ATM and promotes ATM accumulation during HFD-feeding ( a ) Real-time migration of ATM isolated from WAT of lean (chow-fed) or obese (HFD-fed) mice to CCL19 (500 ng/ml). ( b ) Ccr7 mRNA in ATMs isolated from lean or obese WAT. ( c ) Migration of ATM isolated from lean mice to CCL19, netrin-1 (250 ng/ml) or both. ( d ) Migration of ATM isolated from WAT of lean or obese mice to CCL19 in the presence or absence of UNC5b blocking antibody or isotype matched control antibody (Control Ab). ( e–f ) Migration of peritoneal Mø pretreated with 250 µM BSA or palmitate toward CCL19 alone (e), or in the presence of Unc5b-Fc or isotype control (Control-Fc) antibody (f). ( g–h ) body weight ( g ) and fat mass (h) of C57BL6 mice transplanted with WT or Ntn1 −/− bone marrow and fed chow ( n = 7 per group) or HFD ( n = 9 per group) for 20 weeks. ( i ) Emr1 (F4/80) mRNA in VAT of mice of the indicated genotype fed a chow ( n = 4) or western diet ( n = 9). ( j ) Immunofluorescence staining for F4/80 in VAT of representative HFD-fed mice. Scale bar = 100 µm. Data in panels b,d-f are the mean ± s.d. of triplicate samples from a single experiment and are representative of three independent experiments. Data in g-i are the mean ± s.e.m. * P
    Figure Legend Snippet: Netrin-1 blocks chemokine-induced migration of ATM and promotes ATM accumulation during HFD-feeding ( a ) Real-time migration of ATM isolated from WAT of lean (chow-fed) or obese (HFD-fed) mice to CCL19 (500 ng/ml). ( b ) Ccr7 mRNA in ATMs isolated from lean or obese WAT. ( c ) Migration of ATM isolated from lean mice to CCL19, netrin-1 (250 ng/ml) or both. ( d ) Migration of ATM isolated from WAT of lean or obese mice to CCL19 in the presence or absence of UNC5b blocking antibody or isotype matched control antibody (Control Ab). ( e–f ) Migration of peritoneal Mø pretreated with 250 µM BSA or palmitate toward CCL19 alone (e), or in the presence of Unc5b-Fc or isotype control (Control-Fc) antibody (f). ( g–h ) body weight ( g ) and fat mass (h) of C57BL6 mice transplanted with WT or Ntn1 −/− bone marrow and fed chow ( n = 7 per group) or HFD ( n = 9 per group) for 20 weeks. ( i ) Emr1 (F4/80) mRNA in VAT of mice of the indicated genotype fed a chow ( n = 4) or western diet ( n = 9). ( j ) Immunofluorescence staining for F4/80 in VAT of representative HFD-fed mice. Scale bar = 100 µm. Data in panels b,d-f are the mean ± s.d. of triplicate samples from a single experiment and are representative of three independent experiments. Data in g-i are the mean ± s.e.m. * P

    Techniques Used: Migration, Isolation, Mouse Assay, Blocking Assay, Western Blot, Immunofluorescence, Staining

    Netrin-1 expression by ATM promotes metabolic dysfunction ( a ) Serum TNFα levels in mice transplanted with WT or Ntn1 −/− bone marrow and fed chow ( n = 5) or HFD ( n = 6) for 20 wk. ( b ) Glucose tolerance test (GTT) and ( c ) Insulin tolerance test (ITT) of mice fed a chow ( n = 7) or HFD ( n = 9). ( d ) Blood glucose levels and plasma levels of ( e ) insulin, (f) free fatty acid (FFA), and ( g ) adiponectin of mice fed a chow ( n = 7) or HFD ( n = 9). (h) Western blot of phosphoAKT (Ser 473) and total AKT in white adipose tissue (WAT), liver and muscle. Quantification of p-Akt/total Akt is shown at bottom (n=4 mice/group). Data in a-g are the mean ± s.e.m. * P
    Figure Legend Snippet: Netrin-1 expression by ATM promotes metabolic dysfunction ( a ) Serum TNFα levels in mice transplanted with WT or Ntn1 −/− bone marrow and fed chow ( n = 5) or HFD ( n = 6) for 20 wk. ( b ) Glucose tolerance test (GTT) and ( c ) Insulin tolerance test (ITT) of mice fed a chow ( n = 7) or HFD ( n = 9). ( d ) Blood glucose levels and plasma levels of ( e ) insulin, (f) free fatty acid (FFA), and ( g ) adiponectin of mice fed a chow ( n = 7) or HFD ( n = 9). (h) Western blot of phosphoAKT (Ser 473) and total AKT in white adipose tissue (WAT), liver and muscle. Quantification of p-Akt/total Akt is shown at bottom (n=4 mice/group). Data in a-g are the mean ± s.e.m. * P

    Techniques Used: Expressing, Mouse Assay, Western Blot

    Netrin-1 and Unc5b are upregulated in obese-adipose tissue ( a ) Scatter plot of neuronal guidance cue expression in VAT from mice fed a HFD or chow for 20 weeks: upregulated genes are indicated in red; downregulated genes in green ( n = 3 mice/group). Confirmation of mRNA changes in VAT of mice fed chow ( n = 5) or HFD ( n = 6) by qRT-PCR (right panel). ( b ) Western blot of netrin-1, Unc5b, and tubulin in VAT from mice fed chow or HFD ( n = 3). (c) Relative mRNA expression of Ntn1 and Unc5b in the adipocyte and SVF fraction isolated from WAT of mice fed chow ( n = 4) or HFD ( n = 6). ( d ) Immunofluorescence staining for netrin-1 (red, upper) or Unc5b (red, lower), the macrophage marker F4/80 (green) and caveolin-1 (blue) in VAT of HFD-fed mice. Co-localization of netrin-1 or Unc5b with F4/80 is seen in yellow in the merged image (arrows). Scale bar = 100 µm. Data in a and c are the mean ± s.e.m. * P
    Figure Legend Snippet: Netrin-1 and Unc5b are upregulated in obese-adipose tissue ( a ) Scatter plot of neuronal guidance cue expression in VAT from mice fed a HFD or chow for 20 weeks: upregulated genes are indicated in red; downregulated genes in green ( n = 3 mice/group). Confirmation of mRNA changes in VAT of mice fed chow ( n = 5) or HFD ( n = 6) by qRT-PCR (right panel). ( b ) Western blot of netrin-1, Unc5b, and tubulin in VAT from mice fed chow or HFD ( n = 3). (c) Relative mRNA expression of Ntn1 and Unc5b in the adipocyte and SVF fraction isolated from WAT of mice fed chow ( n = 4) or HFD ( n = 6). ( d ) Immunofluorescence staining for netrin-1 (red, upper) or Unc5b (red, lower), the macrophage marker F4/80 (green) and caveolin-1 (blue) in VAT of HFD-fed mice. Co-localization of netrin-1 or Unc5b with F4/80 is seen in yellow in the merged image (arrows). Scale bar = 100 µm. Data in a and c are the mean ± s.e.m. * P

    Techniques Used: Expressing, Mouse Assay, Quantitative RT-PCR, Western Blot, Isolation, Immunofluorescence, Staining, Marker

    3) Product Images from "A Barrel-Related Interneuron in Layer 4 of Rat Somatosensory Cortex with a High Intrabarrel Connectivity"

    Article Title: A Barrel-Related Interneuron in Layer 4 of Rat Somatosensory Cortex with a High Intrabarrel Connectivity

    Journal: Cerebral Cortex (New York, NY)

    doi: 10.1093/cercor/bht263

    Average axonal domains and firing patterns of the 3 different types of fast-spiking L4 interneurons. ( A 1–3) Representative Neurolucida reconstructions of L4 interneurons that were classified as cluster 1 ( A 1), cluster 2 ( A 2), and cluster 3 ( A 3) cells, respectively (axons are given in blue, the somatodendritic domain in red). ( B 1–3) Average axonal length density maps (blue) of the 3 different clusters, respectively. White, thin contour lines enclosing 70%, 80%, and 90% of the integrated axonal density are shown superimposed. The red triangles depict the location of the L4 interneuron somata with respect to the center of the barrel. Note that cluster 1 interneurons have axonal domains that project both throughout cortical layer 2/3, 4, and 5 with the highest density in layer 2/3. In contrast, cluster 3 interneurons have an axonal domain that resides almost exclusively within the home barrel. Cluster 2 interneurons constitute an intermediate class between cluster 1 and 3 neurons with an axon largely confined to the home barrel with only short vertically ascending axonal collaterals in lower layer 2/3. ( C 1–3) Examples of AP firing patterns elicited by rectangular current pulses injected at the soma of the 3 L4 interneuron types. The maximum firing frequency in all of these neurons was ∼300 Hz, that is, all 3 types can be classified as interneurons with FS characteristics. There were no significant differences in the firing characteristics and, thus, the L4 interneurons cannot be discriminated by these criteria. ( D ) Immunohistochemistry of a typical cluster 3 neuron. AMCA streptavidin was used to determine the location and morphology of the neuron, which was positive for PV but negative for CB.
    Figure Legend Snippet: Average axonal domains and firing patterns of the 3 different types of fast-spiking L4 interneurons. ( A 1–3) Representative Neurolucida reconstructions of L4 interneurons that were classified as cluster 1 ( A 1), cluster 2 ( A 2), and cluster 3 ( A 3) cells, respectively (axons are given in blue, the somatodendritic domain in red). ( B 1–3) Average axonal length density maps (blue) of the 3 different clusters, respectively. White, thin contour lines enclosing 70%, 80%, and 90% of the integrated axonal density are shown superimposed. The red triangles depict the location of the L4 interneuron somata with respect to the center of the barrel. Note that cluster 1 interneurons have axonal domains that project both throughout cortical layer 2/3, 4, and 5 with the highest density in layer 2/3. In contrast, cluster 3 interneurons have an axonal domain that resides almost exclusively within the home barrel. Cluster 2 interneurons constitute an intermediate class between cluster 1 and 3 neurons with an axon largely confined to the home barrel with only short vertically ascending axonal collaterals in lower layer 2/3. ( C 1–3) Examples of AP firing patterns elicited by rectangular current pulses injected at the soma of the 3 L4 interneuron types. The maximum firing frequency in all of these neurons was ∼300 Hz, that is, all 3 types can be classified as interneurons with FS characteristics. There were no significant differences in the firing characteristics and, thus, the L4 interneurons cannot be discriminated by these criteria. ( D ) Immunohistochemistry of a typical cluster 3 neuron. AMCA streptavidin was used to determine the location and morphology of the neuron, which was positive for PV but negative for CB.

    Techniques Used: Injection, Immunohistochemistry

    4) Product Images from "Netrin-1 promotes adipose tissue macrophage accumulation and insulin resistance in obesity"

    Article Title: Netrin-1 promotes adipose tissue macrophage accumulation and insulin resistance in obesity

    Journal: Nature medicine

    doi: 10.1038/nm.3467

    Palmitate upregulates Ntn1 and Unc5b expression in macrophages ( a ) Ntn1 and Unc5b mRNA in BMDM treated with 250 µM BSA or palmitate for 24 h. ( b ) Ntn1 and ( c ) Unc5b promoter-luciferase reporter activity in HEK293T cells treated with 250 µM BSA or palmitate in the presence of the NFkB inhibitor BAY11-7082 (10 µmol/L) or vehicle (veh). ( d ) Concentration of netrin-1 in culture supernatants of BMDM treated with 250 µM BSA or palmitate for 24 h in the presence of BAY11-7082 or vehicle. ( e ) Ntn1 and Unc5b mRNA in BMDM treated with conditioned media (CM) from 3T3L1 adipocytes exposed to BSA or palmitate (250 µM) for 24 h. ( f ) qRT-PCR of mRNA from 3T3L1 adipocytes treated with BSA or palmitate (250µM) for 24 h. ( g ) Ntn 1 mRNA in BMDM treated with TNFα (10 ng/ml), IL-6 (40 ng/ml) or IL-4 (10 ng/ml) for 24 h. ( h ) Ntn1 and Unc5b mRNA in BMDM treated with CM from 3T3L1 adipocytes exposed to BSA or palmitate (250 µM) in the presence or absence of anti-TNFα, anti-IL-6 or both. Data presented are the mean ± s.d. of triplicate samples from a single experiment and are representative of three independent experiments. * P
    Figure Legend Snippet: Palmitate upregulates Ntn1 and Unc5b expression in macrophages ( a ) Ntn1 and Unc5b mRNA in BMDM treated with 250 µM BSA or palmitate for 24 h. ( b ) Ntn1 and ( c ) Unc5b promoter-luciferase reporter activity in HEK293T cells treated with 250 µM BSA or palmitate in the presence of the NFkB inhibitor BAY11-7082 (10 µmol/L) or vehicle (veh). ( d ) Concentration of netrin-1 in culture supernatants of BMDM treated with 250 µM BSA or palmitate for 24 h in the presence of BAY11-7082 or vehicle. ( e ) Ntn1 and Unc5b mRNA in BMDM treated with conditioned media (CM) from 3T3L1 adipocytes exposed to BSA or palmitate (250 µM) for 24 h. ( f ) qRT-PCR of mRNA from 3T3L1 adipocytes treated with BSA or palmitate (250µM) for 24 h. ( g ) Ntn 1 mRNA in BMDM treated with TNFα (10 ng/ml), IL-6 (40 ng/ml) or IL-4 (10 ng/ml) for 24 h. ( h ) Ntn1 and Unc5b mRNA in BMDM treated with CM from 3T3L1 adipocytes exposed to BSA or palmitate (250 µM) in the presence or absence of anti-TNFα, anti-IL-6 or both. Data presented are the mean ± s.d. of triplicate samples from a single experiment and are representative of three independent experiments. * P

    Techniques Used: Expressing, Luciferase, Activity Assay, Concentration Assay, Quantitative RT-PCR

    Netrin-1 blocks chemokine-induced migration of ATM and promotes ATM accumulation during HFD-feeding ( a ) Real-time migration of ATM isolated from WAT of lean (chow-fed) or obese (HFD-fed) mice to CCL19 (500 ng/ml). ( b ) Ccr7 mRNA in ATMs isolated from lean or obese WAT. ( c ) Migration of ATM isolated from lean mice to CCL19, netrin-1 (250 ng/ml) or both. ( d ) Migration of ATM isolated from WAT of lean or obese mice to CCL19 in the presence or absence of UNC5b blocking antibody or isotype matched control antibody (Control Ab). ( e–f ) Migration of peritoneal Mø pretreated with 250 µM BSA or palmitate toward CCL19 alone (e), or in the presence of Unc5b-Fc or isotype control (Control-Fc) antibody (f). ( g–h ) body weight ( g ) and fat mass (h) of C57BL6 mice transplanted with WT or Ntn1 −/− bone marrow and fed chow ( n = 7 per group) or HFD ( n = 9 per group) for 20 weeks. ( i ) Emr1 (F4/80) mRNA in VAT of mice of the indicated genotype fed a chow ( n = 4) or western diet ( n = 9). ( j ) Immunofluorescence staining for F4/80 in VAT of representative HFD-fed mice. Scale bar = 100 µm. Data in panels b,d-f are the mean ± s.d. of triplicate samples from a single experiment and are representative of three independent experiments. Data in g-i are the mean ± s.e.m. * P
    Figure Legend Snippet: Netrin-1 blocks chemokine-induced migration of ATM and promotes ATM accumulation during HFD-feeding ( a ) Real-time migration of ATM isolated from WAT of lean (chow-fed) or obese (HFD-fed) mice to CCL19 (500 ng/ml). ( b ) Ccr7 mRNA in ATMs isolated from lean or obese WAT. ( c ) Migration of ATM isolated from lean mice to CCL19, netrin-1 (250 ng/ml) or both. ( d ) Migration of ATM isolated from WAT of lean or obese mice to CCL19 in the presence or absence of UNC5b blocking antibody or isotype matched control antibody (Control Ab). ( e–f ) Migration of peritoneal Mø pretreated with 250 µM BSA or palmitate toward CCL19 alone (e), or in the presence of Unc5b-Fc or isotype control (Control-Fc) antibody (f). ( g–h ) body weight ( g ) and fat mass (h) of C57BL6 mice transplanted with WT or Ntn1 −/− bone marrow and fed chow ( n = 7 per group) or HFD ( n = 9 per group) for 20 weeks. ( i ) Emr1 (F4/80) mRNA in VAT of mice of the indicated genotype fed a chow ( n = 4) or western diet ( n = 9). ( j ) Immunofluorescence staining for F4/80 in VAT of representative HFD-fed mice. Scale bar = 100 µm. Data in panels b,d-f are the mean ± s.d. of triplicate samples from a single experiment and are representative of three independent experiments. Data in g-i are the mean ± s.e.m. * P

    Techniques Used: Migration, Isolation, Mouse Assay, Blocking Assay, Western Blot, Immunofluorescence, Staining

    Netrin-1 and Unc5b are upregulated in obese-adipose tissue ( a ) Scatter plot of neuronal guidance cue expression in VAT from mice fed a HFD or chow for 20 weeks: upregulated genes are indicated in red; downregulated genes in green ( n = 3 mice/group). Confirmation of mRNA changes in VAT of mice fed chow ( n = 5) or HFD ( n = 6) by qRT-PCR (right panel). ( b ) Western blot of netrin-1, Unc5b, and tubulin in VAT from mice fed chow or HFD ( n = 3). (c) Relative mRNA expression of Ntn1 and Unc5b in the adipocyte and SVF fraction isolated from WAT of mice fed chow ( n = 4) or HFD ( n = 6). ( d ) Immunofluorescence staining for netrin-1 (red, upper) or Unc5b (red, lower), the macrophage marker F4/80 (green) and caveolin-1 (blue) in VAT of HFD-fed mice. Co-localization of netrin-1 or Unc5b with F4/80 is seen in yellow in the merged image (arrows). Scale bar = 100 µm. Data in a and c are the mean ± s.e.m. * P
    Figure Legend Snippet: Netrin-1 and Unc5b are upregulated in obese-adipose tissue ( a ) Scatter plot of neuronal guidance cue expression in VAT from mice fed a HFD or chow for 20 weeks: upregulated genes are indicated in red; downregulated genes in green ( n = 3 mice/group). Confirmation of mRNA changes in VAT of mice fed chow ( n = 5) or HFD ( n = 6) by qRT-PCR (right panel). ( b ) Western blot of netrin-1, Unc5b, and tubulin in VAT from mice fed chow or HFD ( n = 3). (c) Relative mRNA expression of Ntn1 and Unc5b in the adipocyte and SVF fraction isolated from WAT of mice fed chow ( n = 4) or HFD ( n = 6). ( d ) Immunofluorescence staining for netrin-1 (red, upper) or Unc5b (red, lower), the macrophage marker F4/80 (green) and caveolin-1 (blue) in VAT of HFD-fed mice. Co-localization of netrin-1 or Unc5b with F4/80 is seen in yellow in the merged image (arrows). Scale bar = 100 µm. Data in a and c are the mean ± s.e.m. * P

    Techniques Used: Expressing, Mouse Assay, Quantitative RT-PCR, Western Blot, Isolation, Immunofluorescence, Staining, Marker

    Related Articles

    Amplification:

    Article Title: Netrin-1 promotes adipose tissue macrophage accumulation and insulin resistance in obesity
    Article Snippet: .. Netrin-1 and Unc5b staining was amplified using the biotin conjugated antibodies (1:100, BA-2000 and BA-9500; Vector Laboratories) followed by streptavidin conjugated AMCA (1:200, Sa-5008; Vector Laboratories) staining. .. Sections were mounted and visualized using a Nikon Eclipse microscope.

    Blocking Assay:

    Article Title: Automated fiber-type-specific cross-sectional area assessment and myonuclei counting in skeletal muscle
    Article Snippet: .. Antibodies (Abs) and reagents used were as follows: anti-dystrophin (1:50; Vector Laboratories, catalog no. VP D505), mouse IgG blocking reagent (catalog no. MKB-2213), streptavidin-AMCA (Alexa Fluor 350; catalog no. SA-5008) all from Vector Laboratories (Burlingame, CA); Texas Red-conjugated goat anti-mouse (catalog no. 610–109-121, Rockland Immunochemicals, Gilbertsville, PA); 4′,6-diamidino-2-phenylindole (DAPI; catalog no. D3571), goat anti-mouse IgG2b, Alexa Fluor 647 conjugated 2° Ab (1:250; catalog no. ), goat anti-mouse IgG1 , Alexa Fluor 488 conjugated 2° Ab (1:500; catalog no. ), goat anti-mouse IgM, biotin conjugated 2° Ab (1:150; catalog no. 626840; Invitrogen, Carlsbad, CA); and anti-MHC I (BA.D5), anti-MHC IIa (SC.71), and anti-MHC IIb (BF.F3) from Developmental Studies Hybridoma Study Bank (Iowa City, IA). ..

    Incubation:

    Article Title: Pkd1 transgenic mice: adult model of polycystic kidney disease with extrarenal and renal phenotypes
    Article Snippet: .. Sections (4 μm) were incubated with one primary antibody α-calbindin (Sigma) for collecting ducts, lycopersicon esculentum lectin (Vector Lab) for distal convoluted tubules and lotus tetragonolobus lectin linked to fluorescein (Vector Lab) for proximal tubules and then with secondary antibodies a goat α-mouse IgG Alexa 255 (Invitrogen) and AMCA-streptavidin (Vector Lab). .. Slides were visualized with Axiovert S100TV microscope.

    other:

    Article Title: A Barrel-Related Interneuron in Layer 4 of Rat Somatosensory Cortex with a High Intrabarrel Connectivity
    Article Snippet: For initial identification of the biocytin-filled neuron, we used AMCA-Streptavidin (Fig. D , upper panel).

    IA:

    Article Title: Automated fiber-type-specific cross-sectional area assessment and myonuclei counting in skeletal muscle
    Article Snippet: .. Antibodies (Abs) and reagents used were as follows: anti-dystrophin (1:50; Vector Laboratories, catalog no. VP D505), mouse IgG blocking reagent (catalog no. MKB-2213), streptavidin-AMCA (Alexa Fluor 350; catalog no. SA-5008) all from Vector Laboratories (Burlingame, CA); Texas Red-conjugated goat anti-mouse (catalog no. 610–109-121, Rockland Immunochemicals, Gilbertsville, PA); 4′,6-diamidino-2-phenylindole (DAPI; catalog no. D3571), goat anti-mouse IgG2b, Alexa Fluor 647 conjugated 2° Ab (1:250; catalog no. ), goat anti-mouse IgG1 , Alexa Fluor 488 conjugated 2° Ab (1:500; catalog no. ), goat anti-mouse IgM, biotin conjugated 2° Ab (1:150; catalog no. 626840; Invitrogen, Carlsbad, CA); and anti-MHC I (BA.D5), anti-MHC IIa (SC.71), and anti-MHC IIb (BF.F3) from Developmental Studies Hybridoma Study Bank (Iowa City, IA). ..

    Staining:

    Article Title: Netrin-1 promotes adipose tissue macrophage accumulation and insulin resistance in obesity
    Article Snippet: .. Netrin-1 and Unc5b staining was amplified using the biotin conjugated antibodies (1:100, BA-2000 and BA-9500; Vector Laboratories) followed by streptavidin conjugated AMCA (1:200, Sa-5008; Vector Laboratories) staining. .. Sections were mounted and visualized using a Nikon Eclipse microscope.

    Binding Assay:

    Article Title: Early intervention with Bifidobacterium lactis NCC2818 modulates the host-microbe interface independent of the sustained changes induced by the neonatal environment
    Article Snippet: .. Binding was detected with isotype specific antisera: goat anti-mouse IgG1 -TRITC; goat anti-mouse IgG2b -FITC; biotinylated rat anti-mouse IgE detected with AMCA-Strepavidin (Vector Laboratories); goat anti-mouse IgG2a AF633 (all Southern Biotechnology). .. Non-specific binding was prevented by 5% pig serum, 5% goat serum and 5% rat serum in PBS.

    Article Title: A Barrel-Related Interneuron in Layer 4 of Rat Somatosensory Cortex with a High Intrabarrel Connectivity
    Article Snippet: .. In addition, we used AMCA-Streptavidin (Vector Laboratories, Burlingame, CA, USA; diluted 1:100) to identify the structural features of the interneuron by its binding to biocytin. .. After incubation with the primary antibodies overnight at 4 °C and subsequent rinsing with Tris-buffered saline (50 mM), the sections were incubated for 2–3 h at room temperature with secondary antibodies, Cy5-conjugated donkey-anti-mouse (1:200; Jackson Laboratories, Bar Harbor, MN, USA) to detect the PV-expression, and Cy3-conjugated goat anti-rabbit (1:200; Vector Laboratories Ltd, Petersborough, UK) to detect the CB-expression in the L4 interneurons, respectively.

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    Vector Laboratories amca streptavidin
    Average axonal domains and firing patterns of the 3 different types of fast-spiking L4 interneurons. ( A 1–3) Representative Neurolucida reconstructions of L4 interneurons that were classified as cluster 1 ( A 1), cluster 2 ( A 2), and cluster 3 ( A 3) cells, respectively (axons are given in blue, the somatodendritic domain in red). ( B 1–3) Average axonal length density maps (blue) of the 3 different clusters, respectively. White, thin contour lines enclosing 70%, 80%, and 90% of the integrated axonal density are shown superimposed. The red triangles depict the location of the L4 interneuron somata with respect to the center of the barrel. Note that cluster 1 interneurons have axonal domains that project both throughout cortical layer 2/3, 4, and 5 with the highest density in layer 2/3. In contrast, cluster 3 interneurons have an axonal domain that resides almost exclusively within the home barrel. Cluster 2 interneurons constitute an intermediate class between cluster 1 and 3 neurons with an axon largely confined to the home barrel with only short vertically ascending axonal collaterals in lower layer 2/3. ( C 1–3) Examples of AP firing patterns elicited by rectangular current pulses injected at the soma of the 3 L4 interneuron types. The maximum firing frequency in all of these neurons was ∼300 Hz, that is, all 3 types can be classified as interneurons with FS characteristics. There were no significant differences in the firing characteristics and, thus, the L4 interneurons cannot be discriminated by these criteria. ( D ) Immunohistochemistry of a typical cluster 3 neuron. <t>AMCA</t> <t>streptavidin</t> was used to determine the location and morphology of the neuron, which was positive for PV but negative for CB.
    Amca Streptavidin, supplied by Vector Laboratories, used in various techniques. Bioz Stars score: 94/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average axonal domains and firing patterns of the 3 different types of fast-spiking L4 interneurons. ( A 1–3) Representative Neurolucida reconstructions of L4 interneurons that were classified as cluster 1 ( A 1), cluster 2 ( A 2), and cluster 3 ( A 3) cells, respectively (axons are given in blue, the somatodendritic domain in red). ( B 1–3) Average axonal length density maps (blue) of the 3 different clusters, respectively. White, thin contour lines enclosing 70%, 80%, and 90% of the integrated axonal density are shown superimposed. The red triangles depict the location of the L4 interneuron somata with respect to the center of the barrel. Note that cluster 1 interneurons have axonal domains that project both throughout cortical layer 2/3, 4, and 5 with the highest density in layer 2/3. In contrast, cluster 3 interneurons have an axonal domain that resides almost exclusively within the home barrel. Cluster 2 interneurons constitute an intermediate class between cluster 1 and 3 neurons with an axon largely confined to the home barrel with only short vertically ascending axonal collaterals in lower layer 2/3. ( C 1–3) Examples of AP firing patterns elicited by rectangular current pulses injected at the soma of the 3 L4 interneuron types. The maximum firing frequency in all of these neurons was ∼300 Hz, that is, all 3 types can be classified as interneurons with FS characteristics. There were no significant differences in the firing characteristics and, thus, the L4 interneurons cannot be discriminated by these criteria. ( D ) Immunohistochemistry of a typical cluster 3 neuron. AMCA streptavidin was used to determine the location and morphology of the neuron, which was positive for PV but negative for CB.

    Journal: Cerebral Cortex (New York, NY)

    Article Title: A Barrel-Related Interneuron in Layer 4 of Rat Somatosensory Cortex with a High Intrabarrel Connectivity

    doi: 10.1093/cercor/bht263

    Figure Lengend Snippet: Average axonal domains and firing patterns of the 3 different types of fast-spiking L4 interneurons. ( A 1–3) Representative Neurolucida reconstructions of L4 interneurons that were classified as cluster 1 ( A 1), cluster 2 ( A 2), and cluster 3 ( A 3) cells, respectively (axons are given in blue, the somatodendritic domain in red). ( B 1–3) Average axonal length density maps (blue) of the 3 different clusters, respectively. White, thin contour lines enclosing 70%, 80%, and 90% of the integrated axonal density are shown superimposed. The red triangles depict the location of the L4 interneuron somata with respect to the center of the barrel. Note that cluster 1 interneurons have axonal domains that project both throughout cortical layer 2/3, 4, and 5 with the highest density in layer 2/3. In contrast, cluster 3 interneurons have an axonal domain that resides almost exclusively within the home barrel. Cluster 2 interneurons constitute an intermediate class between cluster 1 and 3 neurons with an axon largely confined to the home barrel with only short vertically ascending axonal collaterals in lower layer 2/3. ( C 1–3) Examples of AP firing patterns elicited by rectangular current pulses injected at the soma of the 3 L4 interneuron types. The maximum firing frequency in all of these neurons was ∼300 Hz, that is, all 3 types can be classified as interneurons with FS characteristics. There were no significant differences in the firing characteristics and, thus, the L4 interneurons cannot be discriminated by these criteria. ( D ) Immunohistochemistry of a typical cluster 3 neuron. AMCA streptavidin was used to determine the location and morphology of the neuron, which was positive for PV but negative for CB.

    Article Snippet: In addition, we used AMCA-Streptavidin (Vector Laboratories, Burlingame, CA, USA; diluted 1:100) to identify the structural features of the interneuron by its binding to biocytin.

    Techniques: Injection, Immunohistochemistry