Since no universal tissue preparation method will be ideal for all sample and tissue types, the IHC protocol given here is intended as a starting point from which the experimenter should optimize as needed.
Receptor for hyaluronic acid (HA). Mediates cell-cell and cell-matrix interactions through its affinity for HA, and possibly also through its affinity for other ligands such as osteopontin, collagens, and matrix metalloproteinases (MMPs). Adhesion with HA plays an important role in cell migration, tumor growth and progression. In cancer cells, may play an important role in invadopodia formation. Also involved in lymphocyte activation, recirculation and homing, and in hematopoiesis. Altered expression or dysfunction causes numerous pathogenic phenotypes.
Anti-CD44 -http://www.stjohnslabs.com/cd44-antibody-p-91575
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Receptor for hyaluronic acid (HA). Mediates cell-cell and cell-matrix interactions through its affinity for HA, and possibly also through its affinity for other ligands such as osteopontin, collagens, and matrix metalloproteinases (MMPs). Adhesion with HA plays an important role in cell migration, tumor growth and progression. In cancer cells, may play an important role in invadopodia formation. Also involved in lymphocyte activation, recirculation and homing, and in hematopoiesis. Altered expression or dysfunction causes numerous pathogenic phenotypes.
Anti-CD44 -http://www.stjohnslabs.com/cd44-antibody-p-91575
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Pro-inflammatory cytokine. Involved in the innate immune response to bacterial pathogens. The expression of MIF at sites of inflammation suggests a role as mediator in regulating the function of macrophages in host defense. Counteracts the anti-inflammatory activity of glucocorticoids. Has phenylpyruvate tautomerase and dopachrome tautomerase activity (in vitro), but the physiological substrate is not known. It is not clear whether the tautomerase activity has any physiological relevance, and whether it is important for cytokine activity.
Anti-MIF -http://www.stjohnslabs.com/mif-antibody-p-93124
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Toll-like receptor 4 is a protein that in humans is encoded by the TLR4 gene. TLR4 is a transmembrane protein, member of the toll-like receptor family, which belongs to the Pattern Recognition Receptor (PRRs) family. Its activation leads to an intracellular signaling pathway NF-κB and inflammatory cytokine production which is responsible for activating the innate immune system. It is most well known for recognizing lipopolysaccharide (LPS), a component present in many Gram-negative bacteria (e.g. Neisseria spp) and select Gram-positive bacteria. Its ligands also include several viral proteins, polysaccharide, and a variety of endogenous proteins such as low-density lipoprotein, beta-defensins, and heat shock protein.
Anti-CD284 -http://www.stjohnslabs.com/cd284-antibody
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Inhibits the activity of dimeric NF-kappa-B/REL complexes by trapping REL dimers in the cytoplasm through masking of their nuclear localization signals. On cellular stimulation by immune and proinflammatory responses, becomes phosphorylated promoting ubiquitination and degradation, enabling the dimeric RELA to translocate to the nucleus and activate transcription.
Anti-IκB-α -http://www.stjohnslabs.com/ikb-a-antibody
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Plays a key role in the control of the eukaryotic cell cycle by modulating the centrosome cycle as well as mitotic onset; promotes G2-M transition, and regulates G1 progress and G1-S transition via association with multiple interphase cyclins. Required in higher cells for entry into S-phase and mitosis. Essential for early stages of embryonic development. During G2 and early mitosis, CDC25A/B/C-mediated dephosphorylation activates CDK1/cyclin complexes which phosphorylate several substrates that trigger at least centrosome separation, Golgi dynamics, nuclear envelope breakdown and chromosome condensation. Once chromosomes are condensed and aligned at the metaphase plate, CDK1 activity is switched off by WEE1- and PKMYT1-mediated phosphorylation to allow sister chromatid separation, chromosome decondensation, reformation of the nuclear envelope and cytokinesis.
Anti-Cdc2 -http://www.stjohnslabs.com/cdc2-antibody-p-91597
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Tyrosine kinase of the non-receptor type, involved in the IFN-alpha/beta/gamma signal pathway . Kinase partner for the interleukin (IL)-2 receptor. ATP + a [protein]-L-tyrosine = ADP + a [protein]-L-tyrosine phosphate.
Anti-JAK1 -http://www.stjohnslabs.com/jak1-antibody-p-92863
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Calcium-binding protein that interacts with newly synthesized glycoproteins in the endoplasmic reticulum. It may act in assisting protein assembly and/or in the retention within the ER of unassembled protein subunits. It seems to play a major role in the quality control apparatus of the ER by the retention of incorrectly folded proteins. Associated with partial T-cell antigen receptor complexes that escape the ER of immature thymocytes, it may function as a signaling complex regulating thymocyte maturation. Additionally it may play a role in receptor-mediated endocytosis at the synapse.
Anti-Calnexin -http://www.stjohnslabs.com/calnexin-antibody-p-91448
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Transmembrane serine/threonine kinase forming with the TGF-beta type I serine/threonine kinase receptor, TGFBR1, the non-promiscuous receptor for the TGF-beta cytokines TGFB1, TGFB2 and TGFB3. Transduces the TGFB1, TGFB2 and TGFB3 signal from the cell surface to the cytoplasm and is thus regulating a plethora of physiological and pathological processes including cell cycle arrest in epithelial and hematopoietic cells, control of mesenchymal cell proliferation and differentiation, wound healing, extracellular matrix production, immunosuppression and carcinogenesis.
Anti-TGFβ RII -http://www.stjohnslabs.com/tgfb-rii-antibody-p-94590
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Non-receptor protein tyrosine kinase which is activated following engagement of many different classes of cellular receptors including immune response receptors, integrins and other adhesion receptors, receptor protein tyrosine kinases, G protein-coupled receptors as well as cytokine receptors. Participates in signaling pathways that control a diverse spectrum of biological activities including gene transcription, immune response, cell adhesion, cell cycle progression, apoptosis, migration, and transformation. Due to functional redundancy between members of the SRC kinase family, identification of the specific role of each SRC kinase is very difficult. SRC appears to be one of the primary kinases activated following engagement of receptors and plays a role in the activation of other protein tyrosine kinase (PTK) families. Receptor clustering or dimerization leads to recruitment of SRC to the receptor complexes where it phosphorylates the tyrosine residues within the receptor cytoplasmic domains. Plays an important role in the regulation of cytoskeletal organization through phosphorylation of specific substrates such as AFAP1.
Anti-c-Src -http://www.stjohnslabs.com/c-src-antibody
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
In muscle physiology, plays a central role in the balance between atrophy and hypertrophy. When recruited by MSTN, promotes atrophy response via phosphorylated SMAD2/4. MSTN decrease causes SMAD4 release and subsequent recruitment by the BMP pathway to promote hypertrophy via phosphorylated SMAD1/5/8. Acts synergistically with SMAD1 and YY1 in bone morphogenetic protein (BMP)-mediated cardiac-specific gene expression. Binds to SMAD binding elements (SBEs) (5'-GTCT/AGAC-3') within BMP response element (BMPRE) of cardiac activating regions (By similarity). Common SMAD (co-SMAD) is the coactivator and mediator of signal transduction by TGF-beta (transforming growth factor).
Anti-Smad4 -http://www.stjohnslabs.com/smad4-antibody-p-94364
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Serine/threonine-protein kinase that acts downstream of mTOR signaling in response to growth factors and nutrients to promote cell proliferation, cell growth and cell cycle progression. Regulates protein synthesis through phosphorylation of EIF4B, RPS6 and EEF2K, and contributes to cell survival by repressing the pro-apoptotic function of BAD. Under conditions of nutrient depletion, the inactive form associates with the EIF3 translation initiation complex. Upon mitogenic stimulation, phosphorylation by the mammalian target of rapamycin complex 1 (mTORC1) leads to dissociation from the EIF3 complex and activation.
Anti-p70 S6 kinase α -http://www.stjohnslabs.com/p70-s6-kinase-a-antibody-p-93776
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Gap junction protein that acts as a regulator of bladder capacity. A gap junction consists of a cluster of closely packed pairs of transmembrane channels, the connexons, through which materials of low MW diffuse from one cell to a neighboring cell. May play a critical role in the physiology of hearing by participating in the recycling of potassium to the cochlear endolymph. Negative regulator of bladder functional capacity: acts by enhancing intercellular electrical and chemical transmission, thus sensitizing bladder muscles to cholinergic neural stimuli and causing them to contract (By similarity). May play a role in cell growth inhibition through the regulation of NOV expression and localization. Plays an essential role in gap junction communication in the ventricles .
Anti-Connexin 43 -http://www.stjohnslabs.com/connexin-43-antibody-p-91786
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Accelerates programmed cell death by binding to, and antagonizing the apoptosis repressor BCL2 or its adenovirus homolog E1B 19k protein. Under stress conditions, undergoes a conformation change that causes translocation to the mitochondrion membrane, leading to the release of cytochrome c that then triggers apoptosis. Promotes activation of CASP3, and thereby apoptosis.
Anti-Bax -http://www.stjohnslabs.com/bax-antibody-p-91328
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Serine/threonine-protein kinase that acts downstream of mTOR signaling in response to growth factors and nutrients to promote cell proliferation, cell growth and cell cycle progression. Regulates protein synthesis through phosphorylation of EIF4B, RPS6 and EEF2K, and contributes to cell survival by repressing the pro-apoptotic function of BAD. Under conditions of nutrient depletion, the inactive form associates with the EIF3 translation initiation complex. Upon mitogenic stimulation, phosphorylation by the mammalian target of rapamycin complex 1 (mTORC1) leads to dissociation from the EIF3 complex and activation. The active form then phosphorylates and activates several substrates in the pre-initiation complex, including the EIF2B complex and the cap-binding complex component EIF4B. Also controls translation initiation by phosphorylating a negative regulator of EIF4A, PDCD4, targeting it for ubiquitination and subsequent proteolysis.
Anti-p70 S6 kinase α -http://www.stjohnslabs.com/p70-s6-kinase-a-antibody-p-99095
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Promotes cell death. Successfully competes for the binding to Bcl-X(L), Bcl-2 and Bcl-W, thereby affecting the level of heterodimerization of these proteins with BAX. Can reverse the death repressor activity of Bcl-X(L), but not that of Bcl-2 (By similarity). Appears to act as a link between growth factor receptor signaling and the apoptotic pathways.
Anti-Bad -http://www.stjohnslabs.com/bad-antibody-p-91313
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Receptor-regulated SMAD (R-SMAD) that is an intracellular signal transducer and transcriptional modulator activated by TGF-beta (transforming growth factor) and activin type 1 receptor kinases. Binds the TRE element in the promoter region of many genes that are regulated by TGF-beta and, on formation of the SMAD2/SMAD4 complex, activates transcription. May act as a tumor suppressor in colorectal carcinoma. Positively regulates PDPK1 kinase activity by stimulating its dissociation from the 14-3-3 protein YWHAQ which acts as a negative regulator.
Anti-Smad2 -http://www.stjohnslabs.com/smad2-antibody-p-94356
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Probably plays a role in facilitating the assembly of multimeric protein complexes inside the endoplasmic reticulum. Involved in the correct folding of proteins and degradation of misfolded proteins via its interaction with DNAJC10, probably to facilitate the release of DNAJC10 from its substrate
Anti-HSP A5 -http://www.stjohnslabs.com/hsp-a5-antibody
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Pro-inflammatory cytokine. Involved in the innate immune response to bacterial pathogens. The expression of MIF at sites of inflammation suggests a role as mediator in regulating the function of macrophages in host defense. Counteracts the anti-inflammatory activity of glucocorticoids. Has phenylpyruvate tautomerase and dopachrome tautomerase activity (in vitro), but the physiological substrate is not known. It is not clear whether the tautomerase activity has any physiological relevance, and whether it is important for cytokine activity.
Anti-MIF -http://www.stjohnslabs.com/mif-antibody-p-93124
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Toll-like receptor 4 is a protein that in humans is encoded by the TLR4 gene. TLR4 is a transmembrane protein, member of the toll-like receptor family, which belongs to the Pattern Recognition Receptor (PRRs) family. Its activation leads to an intracellular signaling pathway NF-κB and inflammatory cytokine production which is responsible for activating the innate immune system. It is most well known for recognizing lipopolysaccharide (LPS), a component present in many Gram-negative bacteria (e.g. Neisseria spp) and select Gram-positive bacteria. Its ligands also include several viral proteins, polysaccharide, and a variety of endogenous proteins such as low-density lipoprotein, beta-defensins, and heat shock protein.
Anti-CD284 -http://www.stjohnslabs.com/cd284-antibody
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Inhibits the activity of dimeric NF-kappa-B/REL complexes by trapping REL dimers in the cytoplasm through masking of their nuclear localization signals. On cellular stimulation by immune and proinflammatory responses, becomes phosphorylated promoting ubiquitination and degradation, enabling the dimeric RELA to translocate to the nucleus and activate transcription.
Anti-IκB-α -http://www.stjohnslabs.com/ikb-a-antibody
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Plays a key role in the control of the eukaryotic cell cycle by modulating the centrosome cycle as well as mitotic onset; promotes G2-M transition, and regulates G1 progress and G1-S transition via association with multiple interphase cyclins. Required in higher cells for entry into S-phase and mitosis. Essential for early stages of embryonic development. During G2 and early mitosis, CDC25A/B/C-mediated dephosphorylation activates CDK1/cyclin complexes which phosphorylate several substrates that trigger at least centrosome separation, Golgi dynamics, nuclear envelope breakdown and chromosome condensation. Once chromosomes are condensed and aligned at the metaphase plate, CDK1 activity is switched off by WEE1- and PKMYT1-mediated phosphorylation to allow sister chromatid separation, chromosome decondensation, reformation of the nuclear envelope and cytokinesis.
Anti-Cdc2 -http://www.stjohnslabs.com/cdc2-antibody-p-91597
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Tyrosine kinase of the non-receptor type, involved in the IFN-alpha/beta/gamma signal pathway . Kinase partner for the interleukin (IL)-2 receptor. ATP + a [protein]-L-tyrosine = ADP + a [protein]-L-tyrosine phosphate.
Anti-JAK1 -http://www.stjohnslabs.com/jak1-antibody-p-92863
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Calcium-binding protein that interacts with newly synthesized glycoproteins in the endoplasmic reticulum. It may act in assisting protein assembly and/or in the retention within the ER of unassembled protein subunits. It seems to play a major role in the quality control apparatus of the ER by the retention of incorrectly folded proteins. Associated with partial T-cell antigen receptor complexes that escape the ER of immature thymocytes, it may function as a signaling complex regulating thymocyte maturation. Additionally it may play a role in receptor-mediated endocytosis at the synapse.
Anti-Calnexin -http://www.stjohnslabs.com/calnexin-antibody-p-91448
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Transmembrane serine/threonine kinase forming with the TGF-beta type I serine/threonine kinase receptor, TGFBR1, the non-promiscuous receptor for the TGF-beta cytokines TGFB1, TGFB2 and TGFB3. Transduces the TGFB1, TGFB2 and TGFB3 signal from the cell surface to the cytoplasm and is thus regulating a plethora of physiological and pathological processes including cell cycle arrest in epithelial and hematopoietic cells, control of mesenchymal cell proliferation and differentiation, wound healing, extracellular matrix production, immunosuppression and carcinogenesis.
Anti-TGFβ RII -http://www.stjohnslabs.com/tgfb-rii-antibody-p-94590
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Non-receptor protein tyrosine kinase which is activated following engagement of many different classes of cellular receptors including immune response receptors, integrins and other adhesion receptors, receptor protein tyrosine kinases, G protein-coupled receptors as well as cytokine receptors. Participates in signaling pathways that control a diverse spectrum of biological activities including gene transcription, immune response, cell adhesion, cell cycle progression, apoptosis, migration, and transformation. Due to functional redundancy between members of the SRC kinase family, identification of the specific role of each SRC kinase is very difficult. SRC appears to be one of the primary kinases activated following engagement of receptors and plays a role in the activation of other protein tyrosine kinase (PTK) families. Receptor clustering or dimerization leads to recruitment of SRC to the receptor complexes where it phosphorylates the tyrosine residues within the receptor cytoplasmic domains. Plays an important role in the regulation of cytoskeletal organization through phosphorylation of specific substrates such as AFAP1.
Anti-c-Src -http://www.stjohnslabs.com/c-src-antibody
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
In muscle physiology, plays a central role in the balance between atrophy and hypertrophy. When recruited by MSTN, promotes atrophy response via phosphorylated SMAD2/4. MSTN decrease causes SMAD4 release and subsequent recruitment by the BMP pathway to promote hypertrophy via phosphorylated SMAD1/5/8. Acts synergistically with SMAD1 and YY1 in bone morphogenetic protein (BMP)-mediated cardiac-specific gene expression. Binds to SMAD binding elements (SBEs) (5'-GTCT/AGAC-3') within BMP response element (BMPRE) of cardiac activating regions (By similarity). Common SMAD (co-SMAD) is the coactivator and mediator of signal transduction by TGF-beta (transforming growth factor).
Anti-Smad4 -http://www.stjohnslabs.com/smad4-antibody-p-94364
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Serine/threonine-protein kinase that acts downstream of mTOR signaling in response to growth factors and nutrients to promote cell proliferation, cell growth and cell cycle progression. Regulates protein synthesis through phosphorylation of EIF4B, RPS6 and EEF2K, and contributes to cell survival by repressing the pro-apoptotic function of BAD. Under conditions of nutrient depletion, the inactive form associates with the EIF3 translation initiation complex. Upon mitogenic stimulation, phosphorylation by the mammalian target of rapamycin complex 1 (mTORC1) leads to dissociation from the EIF3 complex and activation.
Anti-p70 S6 kinase α -http://www.stjohnslabs.com/p70-s6-kinase-a-antibody-p-93776
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Gap junction protein that acts as a regulator of bladder capacity. A gap junction consists of a cluster of closely packed pairs of transmembrane channels, the connexons, through which materials of low MW diffuse from one cell to a neighboring cell. May play a critical role in the physiology of hearing by participating in the recycling of potassium to the cochlear endolymph. Negative regulator of bladder functional capacity: acts by enhancing intercellular electrical and chemical transmission, thus sensitizing bladder muscles to cholinergic neural stimuli and causing them to contract (By similarity). May play a role in cell growth inhibition through the regulation of NOV expression and localization. Plays an essential role in gap junction communication in the ventricles .
Anti-Connexin 43 -http://www.stjohnslabs.com/connexin-43-antibody-p-91786
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Accelerates programmed cell death by binding to, and antagonizing the apoptosis repressor BCL2 or its adenovirus homolog E1B 19k protein. Under stress conditions, undergoes a conformation change that causes translocation to the mitochondrion membrane, leading to the release of cytochrome c that then triggers apoptosis. Promotes activation of CASP3, and thereby apoptosis.
Anti-Bax -http://www.stjohnslabs.com/bax-antibody-p-91328
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Serine/threonine-protein kinase that acts downstream of mTOR signaling in response to growth factors and nutrients to promote cell proliferation, cell growth and cell cycle progression. Regulates protein synthesis through phosphorylation of EIF4B, RPS6 and EEF2K, and contributes to cell survival by repressing the pro-apoptotic function of BAD. Under conditions of nutrient depletion, the inactive form associates with the EIF3 translation initiation complex. Upon mitogenic stimulation, phosphorylation by the mammalian target of rapamycin complex 1 (mTORC1) leads to dissociation from the EIF3 complex and activation. The active form then phosphorylates and activates several substrates in the pre-initiation complex, including the EIF2B complex and the cap-binding complex component EIF4B. Also controls translation initiation by phosphorylating a negative regulator of EIF4A, PDCD4, targeting it for ubiquitination and subsequent proteolysis.
Anti-p70 S6 kinase α -http://www.stjohnslabs.com/p70-s6-kinase-a-antibody-p-99095
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Promotes cell death. Successfully competes for the binding to Bcl-X(L), Bcl-2 and Bcl-W, thereby affecting the level of heterodimerization of these proteins with BAX. Can reverse the death repressor activity of Bcl-X(L), but not that of Bcl-2 (By similarity). Appears to act as a link between growth factor receptor signaling and the apoptotic pathways.
Anti-Bad -http://www.stjohnslabs.com/bad-antibody-p-91313
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Receptor-regulated SMAD (R-SMAD) that is an intracellular signal transducer and transcriptional modulator activated by TGF-beta (transforming growth factor) and activin type 1 receptor kinases. Binds the TRE element in the promoter region of many genes that are regulated by TGF-beta and, on formation of the SMAD2/SMAD4 complex, activates transcription. May act as a tumor suppressor in colorectal carcinoma. Positively regulates PDPK1 kinase activity by stimulating its dissociation from the 14-3-3 protein YWHAQ which acts as a negative regulator.
Anti-Smad2 -http://www.stjohnslabs.com/smad2-antibody-p-94356
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Probably plays a role in facilitating the assembly of multimeric protein complexes inside the endoplasmic reticulum. Involved in the correct folding of proteins and degradation of misfolded proteins via its interaction with DNAJC10, probably to facilitate the release of DNAJC10 from its substrate
Anti-HSP A5 -http://www.stjohnslabs.com/hsp-a5-antibody
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Sterilisation-
Is the process of making something free from bacteria or other living microorganisms.
Sterility Testing-
Are done to detect if viable forms of micro-organisms are present or not on or in the pharmaceutical preparations.
Involved in the uptake of thrombin-antithrombin complexes by hepatic cells (By similarity). When phosphorylated, plays a role in filament reorganization. Involved in the delivery of mutated CFTR to the plasma membrane. Together with KRT8, is involved in interleukin-6 (IL-6)-mediated barrier protection.
Anti-Cytokeratin 18 -http://www.stjohnslabs.com/cytokeratin-18-antibody-p-91959
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Most upstream protease of the activation cascade of caspases responsible for the TNFRSF6/FAS mediated and TNFRSF1A induced cell death. Binding to the adapter molecule FADD recruits it to either receptor. The resulting aggregate called death-inducing signaling complex (DISC) performs CASP8 proteolytic activation. The active dimeric enzyme is then liberated from the DISC and free to activate downstream apoptotic proteases. Proteolytic fragments of the N-terminal propeptide (termed CAP3, CAP5 and CAP6) are likely retained in the DISC. Cleaves and activates CASP3, CASP4, CASP6, CASP7, CASP9 and CASP10. May participate in the GZMB apoptotic pathways.
Anti-Caspase-8 -http://www.stjohnslabs.com/caspase-8-antibody-p-91485
Caspase-8 Immunofluorescence,Caspase-8 review,CASP8 ,Caspase-8 validation,Caspase-8 report, Anti-Caspase-8 Antibody,Secondary Antibody, antibody report, antibody validation, cell signaling, immunofluorescence, immunofluorescence protocol, immunofluorescence staining, primary antibodies, primary antibody,st john's laboratory, st johns labs,Polyclonal,Rabbit Polyclonal
Anti-Caspase-8 -http://www.stjohnslabs.com/caspase-8-antibody-p-91485
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Complement receptor type 2 (CR2), also known as complement C3d receptor, Epstein-Barr virus receptor, and CD21 (cluster of differentiation 21), is a protein that in humans is encoded by the CR2 gene. CR2 is involved in the complement system. It binds to iC3b (inactive derivative of C3b), C3dg, or C3d. B cells have CR2 receptors on their surfaces, allowing the complement system to play a role in B-cell activation and maturation.
Anti-CD21 -http://www.stjohnslabs.com/cd21-antibody-p-98597
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Serine/threonine kinase which acts as a master kinase, phosphorylating and activating a subgroup of the AGC family of protein kinases. Its targets include: protein kinase B (PKB/AKT1, PKB/AKT2, PKB/AKT3), p70 ribosomal protein S6 kinase (RPS6KB1), p90 ribosomal protein S6 kinase (RPS6KA1, RPS6KA2 and RPS6KA3), cyclic AMP-dependent protein kinase (PRKACA), protein kinase C (PRKCD and PRKCZ), serum and glucocorticoid-inducible kinase (SGK1, SGK2 and SGK3), p21-activated kinase-1 (PAK1), protein kinase PKN (PKN1 and PKN2).
Anti-PDK1 -http://www.stjohnslabs.com/pdk1-antibody-p-93856
Join our Antibody Validation Project - http://www.stjohnslabs.com/services/antibody-validation
Does your next experiment involve Leptin (LEP)? This is a presentation about LEP intended for scientists who are designing controls and performing immunoassays detecting LEP. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-Leptin Antibody Picoband™ (A00479-3):
https://www.bosterbio.com/anti-leptin-picoband-trade-antibody-a00479-3-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC2970652, PMC7520685, PMC3033396
Learn more about LEP (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/LEP
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Fibronectin (FN)? This is a presentation about FN1 intended for scientists who are designing controls and performing immunoassays detecting FN1. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-Fibronectin/FN1 Antibody Picoband™ (A00564-1): https://www.bosterbio.com/anti-fibronectin-fn1-picoband-trade-antibody-a00564-1-boster.html
References: Uniprot.org, ProteinAtlas.org, GeneCards, PMC5112592, PMC3071080,PMC6073216, PMC6204085
Learn more about FN1 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/FN1
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Fibroblast growth factor 23 (FGF23)? This is a presentation about FGF23 intended for scientists who are designing controls and performing immunoassays detecting FGF23. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-FGF23 Antibody Picoband™ (PB9868): https://www.bosterbio.com/anti-fgf23-picoband-trade-antibody-pb9868-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC5595838, PMC7010663, PMC7880350, PMC4121311
Learn more about FGF23 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/FGF23
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Fms-related tyrosine kinase 1 (FLT1)? This is a presentation about FLT1/VEGFR1 intended for scientists who are designing controls and performing immunoassays detecting FLT1/VEGFR1. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-FLT1 Antibody Picoband™ (A00534-4): https://www.bosterbio.com/anti-flt1-picoband-trade-antibody-a00534-4-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC6594043, PMC4710184, PMC2615562
Learn more about FLT1/VEGFR1 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/FLT1
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Plasminogen Activator, Urokinase Receptor (PLAUR)? This is a presentation about uPAR/PLAUR intended for scientists who are designing controls and performing immunoassays detecting uPAR (Urokinase plasminogen activator surface receptor). It contains useful info such as Western blot band size, protein expression, and interesting facts
Anti-UPA Receptor/Plaur Antibody Picoband™ (A00993-2):
https://www.bosterbio.com/anti-upa-receptor-picoband-trade-antibody-a00993-2-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC2586916, PMC6653070, PMC4159539, PMC6355443
Learn more about uPAR/PLAUR (infographic and discussion):
https://www.bosterbio.com/bosterbio-gene-info-cards/PLAUR
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Chemokine (C-X-C motif) ligand 13 (CXCL13)? This is a presentation about CXCL13/BCA1/BLC intended for scientists who are designing controls and performing immunoassays detecting CXCL13/BCA1/BLC. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-BCA1/CXCL13 Antibody Picoband™ (PB9999): https://www.bosterbio.com/anti-bca1-picoband-trade-antibody-pb9999-boster.html
References: Uniprot.org, ProteinAtlas.org, GeneCards, PMC5307320, PMC7422843, PMC4940825
Learn more about CXCL13 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/CXCL13
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Integrin alpha-3? This is a presentation about Integrin alpha 3/ITGA3 intended for scientists who are designing controls and performing immunoassays detecting Integrin alpha 3/ITGA3. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-Integrin Alpha 3/ITGA3 Antibody (PA1621):
https://www.bosterbio.com/anti-integrin-alpha-3-antibody-pa1621-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC6034741, PMC2947217, PMC7327512
Learn more about Integrin alpha 3/ITGA3 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/ITGA3
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Chemokine (C-C motif) ligand 2 (CCL2)? This is a presentation about MCP-1/CCL2 intended for scientists who are designing controls and performing immunoassays detecting MCP-1/CCL2. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-MCP1/CCL2 Antibody Picoband™ (PB9570): https://www.bosterbio.com/anti-mcp-1-picoband-trade-antibody-pb9570-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC5261840, PMC8854715, PMC6883042, PMC5428676
Learn more about MCP-1/CCL2 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/CCL2
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Insulin-like growth factor I (IGF-I, IGF-1)? This is a presentation about IGF1 intended for scientists who are designing controls and performing immunoassays detecting IGF1. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-IGF1 Mouse Monoclonal Antibody (M00148-1): https://www.bosterbio.com/anti-igf1-mouse-monoclonal-antibody-clone-id-oti4b12-m00148-1-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC5867890, PMC7450330, PMC7262660, PMC7401641
Learn more about IGF1 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/IGF1
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Periostin (POSTN)? This is a presentation about POSTN intended for scientists who are designing controls and performing immunoassays detecting POSTN. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-Periostin/Postn Antibody Picoband™ (A01378):
https://www.bosterbio.com/anti-periostin-picoband-trade-antibody-a01378-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC4826188, PMC4746667, PMC4262539, PMC3443161
Learn more about POSTN (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/POSTN
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Angiotensin-converting enzyme (ACE)? This is a presentation about ACE intended for scientists who are designing controls and performing immunoassays detecting ACE. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-Angiotensin Converting Enzyme 1/ACE Antibody (PA2196-2): https://www.bosterbio.com/anti-angiotensin-converting-enzyme-1-antibody-pa2196-2-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC3322615, PMC5722543, PMC6672927, PMC3389005
Learn more about ACE (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/ACE
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Lipopolysaccharide binding protein (LBP)? This is a presentation about LBP intended for scientists who are designing controls and performing immunoassays detecting LBP. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-LBP Antibody Picoband™ (A00809-1):
https://www.bosterbio.com/anti-lbp-picoband-trade-antibody-a00809-1-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC3617275, PMC3485520, PMC6945416, PMC8710911
Learn more about LBP (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/LBP
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve NGFβ? This is a presentation about NGF/NGF Beta intended for scientists who are designing controls and performing immunoassays detecting NGF/beta-NGF. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-NGF/NGF Beta Antibody Picoband™ (A00341): https://www.bosterbio.com/anti-ngf-ngf-beta-picoband-trade-antibody-a00341-boster.html
References: Uniprot.org, ProteinAtlas.org, GeneCards, PMC4760111, PMC6037903, PMC2650228, PMC1894684
Learn more about NGF/NGF Beta (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/NGF
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve CD133 (Prominin-1)? This is a presentation about CD133/Prominin-1/PROM1 intended for scientists who are designing controls and performing immunoassays detecting Prominin-1. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-PROM1 Antibody Picoband™ (PB9156):
https://www.bosterbio.com/anti-prom1-picoband-trade-antibody-pb9156-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC6763460, PMC2713505, PMC6030467
Learn more about CD133/Prominin-1/PROM1 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/PROM1
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve CSF3? This is a presentation about CSF3/G-CSF intended for scientists who are designing controls and performing immunoassays detecting CSF3/G-CSF. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-G-CSF CSF3 Rabbit Monoclonal Antibody (M02280-1):
https://www.bosterbio.com/anti-g-csf-rabbit-monoclonal-antibody-m02280-1-boster.html
Anti-G-CSF/CSF3 Antibody Picoband™ (PB9563):
https://www.bosterbio.com/anti-g-csf-picoband-trade-antibody-pb9563-boster.html
References: Uniprot.org, ProteinAtlas.org, GeneCards, PMC5758815, PMC7196588, PMC8086136
Learn more about CSF3/G-CSF (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/CSF3
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve RANKL? This is a presentation about RANKL/TRANCE intended for scientists who are designing controls and performing immunoassays detecting TNFSF11/RANKL/TRANCE. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-RANKL/TNFSF11 Antibody Picoband™ (PB10015):
https://www.bosterbio.com/anti-rankl-picoband-trade-antibody-pb10015-boster.html
References: Uniprot.org, ProteinAtlas.org, GeneCards, PMC8602693, PMID 29203513, PMC7949628
Learn more about TNFSF11/RANKL/TRANCE/OPGL/ODF (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/TNFSF11
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Neurofilament Heavy Polypeptide (NEFH)? This is a presentation about NEFH/NF-H intended for scientists who are designing controls and performing immunoassays detecting NEFH/NF-H. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-NEFH/Nf H Rabbit Monoclonal Antibody (M05307-2):
https://www.bosterbio.com/anti-nefh-rabbit-monoclonal-antibody-m05307-2-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC6834541, PMC6054723, PMC4285109
Learn more about NEFH/NF-H (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/NEFH
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Kidney injury molecule 1 (KIM-1)? This is a presentation about KIM1/TIM1/HAVCR1 intended for scientists who are designing controls and performing immunoassays detecting KIM-1/TIM-1/HAVCR1. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-TIM 1/HAVCR1 Antibody (PA1624): https://www.bosterbio.com/anti-tim-1-antibody-pa1624-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC4552842, PMC8134587, PMC8139578, PMC7414978
Learn more about KIM-1/TIM-1/HAVCR1 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/HAVCR1
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Interferon gamma? This is a presentation about IFN Gamma/IFNG intended for scientists who are designing controls and performing immunoassays detecting IFN Gamma/IFNG. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-Interferon Gamma IFNG Rabbit Monoclonal Antibody (M00393-1): https://www.bosterbio.com/anti-interferon-gamma-rabbit-monoclonal-antibody-m00393-1-boster.html
Anti-IFN Gamma Antibody (RP1001):
https://www.bosterbio.com/anti-rat-ifn-gamma-antibody-rp1001-boster.html
References: Uniprot.org, ProteinAtlas.org, PMID15220936, PMID9524237, PMID24253448, PMC7086207
Learn more about IFN Gamma/IFNG (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/IFNG
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Does your next experiment involve Bone morphogenetic protein 2 (BMP-2)? This is a presentation about BMP-2 intended for scientists who are designing controls and performing immunoassays detecting BMP-2. It contains useful info such as Western blot band size, protein expression, and interesting facts.
Anti-BMP2 Rabbit Polyclonal Antibody (A00338): https://www.bosterbio.com/anti-bmp2-rabbit-polyclonal-antibody-a00338-boster.html
References: Uniprot.org, ProteinAtlas.org, PMC5645599, PMC8082363, PMC5425130, PMC8170469
Learn more about BMP-2 (infographic and discussion): https://www.bosterbio.com/bosterbio-gene-info-cards/BMP2
Boster Biological Technology
Website: www.bosterbio.com
Email: support@bosterbio.com
Seminar of U.V. Spectroscopy by SAMIR PANDASAMIR PANDA
Spectroscopy is a branch of science dealing the study of interaction of electromagnetic radiation with matter.
Ultraviolet-visible spectroscopy refers to absorption spectroscopy or reflect spectroscopy in the UV-VIS spectral region.
Ultraviolet-visible spectroscopy is an analytical method that can measure the amount of light received by the analyte.
A brief information about the SCOP protein database used in bioinformatics.
The Structural Classification of Proteins (SCOP) database is a comprehensive and authoritative resource for the structural and evolutionary relationships of proteins. It provides a detailed and curated classification of protein structures, grouping them into families, superfamilies, and folds based on their structural and sequence similarities.
This presentation explores a brief idea about the structural and functional attributes of nucleotides, the structure and function of genetic materials along with the impact of UV rays and pH upon them.
Observation of Io’s Resurfacing via Plume Deposition Using Ground-based Adapt...Sérgio Sacani
Since volcanic activity was first discovered on Io from Voyager images in 1979, changes
on Io’s surface have been monitored from both spacecraft and ground-based telescopes.
Here, we present the highest spatial resolution images of Io ever obtained from a groundbased telescope. These images, acquired by the SHARK-VIS instrument on the Large
Binocular Telescope, show evidence of a major resurfacing event on Io’s trailing hemisphere. When compared to the most recent spacecraft images, the SHARK-VIS images
show that a plume deposit from a powerful eruption at Pillan Patera has covered part
of the long-lived Pele plume deposit. Although this type of resurfacing event may be common on Io, few have been detected due to the rarity of spacecraft visits and the previously low spatial resolution available from Earth-based telescopes. The SHARK-VIS instrument ushers in a new era of high resolution imaging of Io’s surface using adaptive
optics at visible wavelengths.
Slide 1: Title Slide
Extrachromosomal Inheritance
Slide 2: Introduction to Extrachromosomal Inheritance
Definition: Extrachromosomal inheritance refers to the transmission of genetic material that is not found within the nucleus.
Key Components: Involves genes located in mitochondria, chloroplasts, and plasmids.
Slide 3: Mitochondrial Inheritance
Mitochondria: Organelles responsible for energy production.
Mitochondrial DNA (mtDNA): Circular DNA molecule found in mitochondria.
Inheritance Pattern: Maternally inherited, meaning it is passed from mothers to all their offspring.
Diseases: Examples include Leber’s hereditary optic neuropathy (LHON) and mitochondrial myopathy.
Slide 4: Chloroplast Inheritance
Chloroplasts: Organelles responsible for photosynthesis in plants.
Chloroplast DNA (cpDNA): Circular DNA molecule found in chloroplasts.
Inheritance Pattern: Often maternally inherited in most plants, but can vary in some species.
Examples: Variegation in plants, where leaf color patterns are determined by chloroplast DNA.
Slide 5: Plasmid Inheritance
Plasmids: Small, circular DNA molecules found in bacteria and some eukaryotes.
Features: Can carry antibiotic resistance genes and can be transferred between cells through processes like conjugation.
Significance: Important in biotechnology for gene cloning and genetic engineering.
Slide 6: Mechanisms of Extrachromosomal Inheritance
Non-Mendelian Patterns: Do not follow Mendel’s laws of inheritance.
Cytoplasmic Segregation: During cell division, organelles like mitochondria and chloroplasts are randomly distributed to daughter cells.
Heteroplasmy: Presence of more than one type of organellar genome within a cell, leading to variation in expression.
Slide 7: Examples of Extrachromosomal Inheritance
Four O’clock Plant (Mirabilis jalapa): Shows variegated leaves due to different cpDNA in leaf cells.
Petite Mutants in Yeast: Result from mutations in mitochondrial DNA affecting respiration.
Slide 8: Importance of Extrachromosomal Inheritance
Evolution: Provides insight into the evolution of eukaryotic cells.
Medicine: Understanding mitochondrial inheritance helps in diagnosing and treating mitochondrial diseases.
Agriculture: Chloroplast inheritance can be used in plant breeding and genetic modification.
Slide 9: Recent Research and Advances
Gene Editing: Techniques like CRISPR-Cas9 are being used to edit mitochondrial and chloroplast DNA.
Therapies: Development of mitochondrial replacement therapy (MRT) for preventing mitochondrial diseases.
Slide 10: Conclusion
Summary: Extrachromosomal inheritance involves the transmission of genetic material outside the nucleus and plays a crucial role in genetics, medicine, and biotechnology.
Future Directions: Continued research and technological advancements hold promise for new treatments and applications.
Slide 11: Questions and Discussion
Invite Audience: Open the floor for any questions or further discussion on the topic.
(May 29th, 2024) Advancements in Intravital Microscopy- Insights for Preclini...Scintica Instrumentation
Intravital microscopy (IVM) is a powerful tool utilized to study cellular behavior over time and space in vivo. Much of our understanding of cell biology has been accomplished using various in vitro and ex vivo methods; however, these studies do not necessarily reflect the natural dynamics of biological processes. Unlike traditional cell culture or fixed tissue imaging, IVM allows for the ultra-fast high-resolution imaging of cellular processes over time and space and were studied in its natural environment. Real-time visualization of biological processes in the context of an intact organism helps maintain physiological relevance and provide insights into the progression of disease, response to treatments or developmental processes.
In this webinar we give an overview of advanced applications of the IVM system in preclinical research. IVIM technology is a provider of all-in-one intravital microscopy systems and solutions optimized for in vivo imaging of live animal models at sub-micron resolution. The system’s unique features and user-friendly software enables researchers to probe fast dynamic biological processes such as immune cell tracking, cell-cell interaction as well as vascularization and tumor metastasis with exceptional detail. This webinar will also give an overview of IVM being utilized in drug development, offering a view into the intricate interaction between drugs/nanoparticles and tissues in vivo and allows for the evaluation of therapeutic intervention in a variety of tissues and organs. This interdisciplinary collaboration continues to drive the advancements of novel therapeutic strategies.
2. Recommended Protocols
Tissue preparation is a key to successful IHC experiments. Since no universal tissue preparation method will be ideal for all
sample and tissue types, all protocols given here are intended as a starting point from which the experimenter must optimize as
needed. All conditions should be standardized in order to ensure reproducible results. Keep in mind that you must be careful not
to allow tissues to dry out at any time.
1. IHC (ParaffinSections)
Figure 7. IHC (Paraffin Sections) Workflow with Applicable Boster’s Reagents.
(A) Tissue Preparation
(i) Paraformaldehyde Cooling and Dehydration
Harvest fresh tissue and place it in a dish filled with ice-cold PBS buffer
Wash the tissue thoroughly with PBS to remove blood (Use forceps to remove connective tissues)
Cut the tissue into slices of thickness of 3 mm or less
3. Immerse the slices in 4% paraformaldehyde at room temperature for 8 min
Immerse the slices in 4% paraformaldehyde (pre-cool at 4°C) for 6 to 7 hrs. The paraformaldehyde volume
should be 20X greater than the tissue volume by weight
Wash the tissue 3X with PBS (1 min each)
Dehydrate the tissue by immersing the tissue sequentially as follows:
- 1X into 80% ethanol (1 hr at4°C)
- 1X into 90% ethanol (1 hr at4°C)
- 3X into 95% ethanol (1 hr each at 4°C)
- 3X into 100% ethanol (1 hr each at 4°C)
- 3X into dimethylbenzene (0.5 hr each at room temperature)
25
(ii) Liquid Paraffin Section
Prepare the first portion of liquid paraffin in a suitable bath and allow the paraffin to reach and maintain at
60°C
Immerse the tissue 2X into the paraffin bath (2 hrseach)
Prepare the second portion of liquid paraffin in a suitable bath and allow the paraffin to reach and maintain at
60°C
Pour the second portion of paraffin into a mold
Quickly transport the tissue from the paraffin bath to the mold with paraffin
Incubate the tissue at room temperature until it coagulates
Store the tissue at 4°C
(iii) Section Slicing and Incubation
Secure the paraffin section on slicer
Slice one to two pieces of section to adjust the slicer so that the section and blade are parallel
Slice the remaining section carefully with ~5 µm thick
Incubate the sliced section in 40 to 50°C water to unfold
Mount the tissue section onto Poly-Lysine or APES coated glass slides
Incubate the slides overnight at 37°C
4. (B) Dewaxing/Deparaffinization
Prepare the following reagents:
- 90% dimethylbenzene
- 95% dimethylbenzene
- 100% dimethylbenzene
- 90%ethanol
- 95%ethanol
- 100% ethanol
Sequentially immerse paraffin sections into:
- 90% dimethylbenzene (for 7 min)
- 95% dimethylbenzene (for 7 min)
- 100% dimethylbenzene (for 7 min)
- 90% ethanol (for 7 min)
- 95% ethanol (for 7 min)
- 100% ethanol (for 7 min)
Wash the slides with water to remove ethanol
Note: The process of dewaxing should be done in a fume hood at room temperature in summer. When the
temperature is lower than 18°C, it is recommended to dewax at 50°C.
(C) Inactivation
Immerse dewaxed paraffin section into the 3% H2O2 at room temperature for 10min
Wash the section 3X to 5X with distilled water (total 3 to 5 min)
(D) Antigen Retrieval (Heat Induced Epitope Retrieval: HIER)
Immerse the paraffin sections in citrate buffer
Heat the buffer in microwave and turn it off when the buffer has boiled
Keep the boiled buffer in microwave for 5 to 10 min
Repeat the heating as outlined above 1X to 2X
Cool the slides until it reaches room temperature
Wash the sections 1X to 2X with PBS
5. (C) Blocking
Add 5% BSA blocking solution or normal goat serum to the HIER treated samples
Incubate the samples at 37°C for 30 min
Discard extra liquid (No washing required)
(D) Primary Antibody Incubation
Dilute primary antibody with antibody diluent to the concentration recommended by the antibody manufacturer
Add the diluted antibody to the samples and incubate at 37°C overnight
Wash the samples 2X with PBS (20 min each)
(E) Secondary Antibody Incubation
Dilute biotinylated secondary antibody with antibody diluent to the concentration recommended by the antibody manufacturer
Add the diluted antibody to the samples and incubate at 37°C for 30 min
Wash the samples 2X with PBS (20 min each)
(F) Staining
Add Strept-Avidin Biotin Complex (SABC) HRP- or AP-conjugated reagents to the samples
Incubate the samples at 37°C for 30 min
Wash the samples 3X with PBS (20 min each)
Add a suitable amount of DAB reagent to the samples and incubate in dark at room temperature for 10 to 30 min
Monitor the tissue staining intensity under a bright-field microscope*
Wash the samples 3X to 5X with distilled water
Counterstain (if necessary)
- Add haematoxylin to the sample
- Dehydrate
- Immerse the paraffin sections 2X in dimethylbenzene (7 min each)
Check the tissue staining intensity under a bright-field microscope
* If the staining background is too high, wash the section 4X with 0.01-0.02% TWEEN 20 PBS and 2X with pure PBS after the SABC reaction and before DAB
staining. Then use DAB to stain the samples.
6. 2. IHC (Frozen Sections)
Figure 8. IHC (Frozen Sections) Workflow with Applicable Boster’s Reagents.
(A) Tissue Preparation
(i) Snap Freezing and OCT Embedding
Harvest fresh tissue and place it in a dish filled with ice-cold PBS buffer
Wash the tissue thoroughly with PBS to remove blood (Use forceps to remove connective tissues)
Cut the tissue into slices of thickness of 3 mm or less
Immediately snap freeze the tissue in iso-pentane cooled in dry ice and keep the tissue at
-70°C (Do not allow frozen tissue to thaw beforecutting)
Prior to cryostat sectioning, position the tissue in a mold (which can be simply made by using tin foil) and cover
the tissue completely in Optimal Cutting Temperature (OCT) embedding medium
Use forceps to take the bottom part of mold into liquid nitrogen for 1 to 2 min (The OCT should change to
white)
(i) Cryostat Sectioning
Pre-cool a slicer box and detector to -22°C and -24°C, respectively (Ensure the completeness and smoothness
of blade)
Place the tissue from the mold to the detector where the tissue is fixed
7. Quickly and carefully slice the cryostat sections at 5-10 µm and mount them on gelatin-coated histological slides. Note that:
- Use coverslip to take sliced tissue
- Cryostat temperature should be between -15°C and -23°C
- The sections will curl up if the specimen is too cold
- The sections will stick to the knife if the specimen is too warm
Air dry the sections at room temperature for 30 min to prevent them from falling off the slides during antibody incubations
Store the slides at -70°C. Note that:
- The slides can be stored unfixed for several months at -70°C
- Frozen tissue samples saved for later analysis should be stored intact
Immediately add 50 µL of ice-cold fixation buffer to each tissuesection upon removal from the freezer
Fix frozen section by immersing it into 4% paraformaldehyde at 2-8°C for 8 min (Or optimally at -20°C for 20 min)
Wash the section 3X with PBS and allow it to dry at room temperature for 30 min
(B) Inactivation
• Immerse dewaxed paraffin section into the 3% H2O2 at room temperature for 10 min
• Wash the section 3X to 5X with distilled water (total 3 to 5 min)
(C) Blocking
Add 5% BSA blocking solution or normal goat serum to the HIER treated samples
Incubate the samples at 37°C for 30 min
Discard extra liquid (No washing required)
D) Antigen Retrieval (Heat Induced Epitope Retrieval: HIER)
• Immerse the paraffin sections in citrate buffer
• Heat the buffer in microwave and turn it off when the buffer has boiled
• Keep the boiled buffer in microwave for 5 to 10 min
• Repeat the heating as outlined above 1X to 2X
• Cool the slides until it reaches room temperature
• Wash the sections 1X to 2X with PBS
2
8.
31
Add the diluted antibody to the samples and incubate at 37°C for 30 min Wash the
samples 2X with PBS (20 min each)
(G) Staining
Add Strept-Avidin Biotin Complex (SABC) HRP- or AP-conjugated reagents to the samples Incubate the
samples at 37°C for 30 min
Wash the samples 3X with PBS (20 min each)
Add a suitable amount of DAB reagent to the samples and incubate in dark at room temperature for 10 to 30 min
Monitor the tissue staining intensity under a bright-field microscope* Wash the samples
3X to 5X with distilled water
Counterstain (if necessary)
- Add haematoxylin to the sample
- Dehydrate
- Immerse the paraffin sections 2X in dimethylbenzene (7 min each)
Check the tissue staining intensity under a bright-field microscope
* If the staining background is too high, wash the section 4X with 0.01-0.02% TWEEN 20 PBS and 2X with pure PBS
after the SABC reaction and before DAB staining. Then use DAB to stain the samples.
(E) Primary Antibody Incubation
Dilute primary antibody with antibody diluent to the concentration recommended by the antibody
manufacturer
Add the diluted antibody to the samples and incubate at 37°C overnight
Wash the samples 2X with PBS (20 min each)
(F) Secondary Antibody Incubation
Dilute biotinylated secondary antibody with antibody diluent o the concentration recommended by
the antibody manufacturer
9. 3. ICC/IF (Cell Climbing Slices)
Figure 9. ICC/IF Workflow with Applicable Boster’s Reagents.
30
(A) Cell Climbing Slice Preparation
Place settled coverslip in culture bottle or perforated plate
Take out coverslip after cell growth has reached 60%
Wash the coverslip 3X with PBS to remove culture medium
Immerse the coverslip (cells face up) into cold acetone or 4% paraformaldehyde or neutral
formalin for 10 to 20 min (Close the lid to prevent evaporation)
Wash the coverslip 3X with PBS
Put the coverslip on filter paper (cells face up)
Remove the liquid on the coverslip and allow it to dry for 8-10 hrs
To thaw the slice, wash with neutral PBS at room temperature for 10-15 min (The cell
climbing slice can be stored in gelatin at -20°C for one week.)
10. Note: This fixation procedure using paraformaldehyde and formalin fixatives may cause autofluorescence in the
green spectrum. In this case, you may try fluorophores in the (i) red range or (ii) infrared range if you have an
infrared detection system.
(B) Inactivation
Mix H2O2 with distilled water (v/v: 1:50)
Immerse frozen section or cell climbing slice into the diluted H2O2 at room temperature for 10 min
Wash the section 3X distilled water (1 min each)
(C) Antigen Retrieval (Proteolytic Induced Epitope Retrieval: PIER)
Dry the cell slices with filter paper
Add compound digestion solution (e.g. Trypsin solution or other enzymatic antigen retrieval solution) to the
slices (We recommend the addition of 0.1% Triton to the samples before the digestion. This reduces surface
tension and allows reagents to easily cover the entire sample.)
Incubate the slices at room temperature for 10 min
Wash with 3X PBS (10 min each)
(D) Blocking
Add 5% BSA blocking solution or normal goat serum to the PIER treated samples
Incubate the samples at 37°C for 30 min
Shake off extra liquid and dry the samples with filter paper (No washing required)
(E) Primary Antibody Incubation
Dilute primary antibody with antibody diluent to the concentration recommended by the antibody manufacturer
Add the diluted antibody (Recommended concentration: 0.4 μg to 2 μg) to the samples and incubate at 4°C
overnight
Wash the samples 3X with PBS (15 min each)
11. (F) Secondary Antibody Incubation
Dilute biotinylated secondary antibody with antibody diluent to the concentration recommended by the antibody
manufacturer
Add the diluted antibody to the samples and incubate at 37°C for 30 min
Wash the samples 3X with PBS (8 min each)
(G) Staining
Add Strept-Avidin Biotin Complex – Fluorescence Iso-Thio-Cyanate (SABC-FITC) or Strept-Avidin Biotin Complex
– Cyanine-3 (SABC-Cy3) reagents to the samples
Incubate the samples at 37°C for 30 min (Avoid light)
Wash the samples 2X with PBS (Total 2 hrs)
Seal the slices with water soluble sealing reagent
Monitor the staining intensity under a fluorescence microscope
Counterstain by adding DAPI staining solution to the sample
Check again the staining intensity under a fluorescence microscope
For slide storage without significant decay in fluorescence signal, add 20 μL of anti-fade solution to the sample
followed by a cover glass (Avoid bubbles)