HSD1 Antibody

What is 11β-HSD1 and why is it significant in research?

11β-HSD1 (Corticosteroid 11-beta-dehydrogenase isozyme 1) is a microsomal enzyme that primarily functions as a NADP(H)-dependent reductase, converting inactive cortisone to active cortisol, thereby regulating intracellular cortisol access to glucocorticoid receptors . This enzyme is widely expressed in liver and adipose tissue, as well as adrenal gland, ovary, and decidua . It plays a critical role in local glucocorticoid metabolism and has been implicated in various pathologies including obesity, metabolic syndrome, and inflammatory conditions . The bidirectional nature of 11β-HSD1 activity (both reductase and dehydrogenase) makes it an important target for studying glucocorticoid regulation in different tissue microenvironments .

What are the common alternative names for HSD11B1?

The enzyme is known by several alternative names in the scientific literature:

These alternative names appear frequently in publications and database entries, so researchers should be aware of all nomenclature when conducting literature searches or database mining .

What is the molecular weight discrepancy in 11β-HSD1 detection?

A notable characteristic of 11β-HSD1 is the difference between its calculated and observed molecular weights. While the calculated molecular weight is approximately 36 kDa, many researchers observe bands at 65-70 kDa in Western blot analyses . This discrepancy is due to post-translational modifications, particularly glycosylation of the native protein. Using appropriate positive controls (such as liver tissue extracts) is essential when validating antibody specificity, as this discrepancy could otherwise be mistakenly interpreted as non-specific binding .

What are the recommended applications and dilutions for 11β-HSD1 antibodies?

11β-HSD1 antibodies have been validated for multiple experimental applications with specific dilution ranges:

It is strongly recommended that researchers titrate the antibody in each specific testing system to obtain optimal results, as the appropriate dilution may be sample-dependent .

How should 11β-HSD1 antibodies be stored for optimal performance?

For maximum stability and activity retention, 11β-HSD1 antibodies should be stored at -20°C . Most commercial preparations remain stable for one year after shipment when maintained under these conditions. Many commercially available antibodies are supplied in PBS with 0.02% sodium azide and 50% glycerol (pH 7.3), which helps maintain stability . Aliquoting is generally unnecessary for -20°C storage, particularly for smaller volume preparations (e.g., 20μl sizes that contain 0.1% BSA) .

What positive controls are recommended for validating 11β-HSD1 antibodies?

Based on documented expression patterns, the following samples serve as reliable positive controls:

These controls have been validated in multiple studies and represent tissues/cells with confirmed endogenous expression of 11β-HSD1 .

What is the tissue distribution pattern of 11β-HSD1?

11β-HSD1 shows a distinctive tissue expression pattern that should be considered when designing experiments:

This expression profile helps researchers anticipate where 11β-HSD1 can be reliably detected and what tissues might serve as appropriate positive or negative controls .

How is 11β-HSD1 expressed in immune cells, particularly during inflammation?

11β-HSD1 is dynamically expressed in immune cells, with particularly notable expression in neutrophils during inflammatory responses . During thioglycollate-induced peritonitis in mouse models, neutrophils (identified as CD11b+, Ly6G+, 7/4+ cells) show high expression of 11β-HSD1 early in inflammation, with expression decreasing at later stages . This temporal regulation differs between tissue-localized and circulating neutrophils; peritoneal neutrophils show declining 11β-HSD1 expression as inflammation resolves, while blood neutrophils continue to increase expression during the inflammatory process .

When designing experiments to investigate 11β-HSD1 in inflammatory conditions, researchers should consider:

• Time course sampling is essential due to dynamic regulation

• Different compartments (blood vs. tissue) show distinct expression patterns

• Cell-specific markers should be used to differentiate neutrophil populations from other myeloid cells

How does the relationship between H6PDH and 11β-HSD1 affect experimental outcomes?

Hexose-6-phosphate dehydrogenase (H6PDH) plays a crucial role in determining the directionality of 11β-HSD1 enzymatic activity . Within the endoplasmic reticulum, H6PDH generates NADPH, which drives 11β-HSD1 to function predominantly as a reductase (converting inactive to active glucocorticoids) . In the absence of sufficient NADPH or H6PDH, 11β-HSD1 can switch to functioning as a dehydrogenase, inactivating glucocorticoids similar to 11β-HSD2 .

This relationship has significant experimental implications:

• When studying 11β-HSD1 activity in cell-free systems, inclusion of NADPH is necessary to observe reductase activity

• In tissues with low H6PDH expression, 11β-HSD1 may primarily function as a dehydrogenase

• Co-expression studies of both enzymes may be necessary to fully understand glucocorticoid metabolism in specific tissues

How can genetic or pharmacological modulation of 11β-HSD1 inform inflammatory research?

Both genetic deletion and pharmacological inhibition of 11β-HSD1 have been shown to augment inflammatory cell recruitment during experimental peritonitis . Specifically, 11β-HSD1-deficient mice demonstrate enhanced recruitment of inflammatory cells and delayed acquisition of macrophage phagocytic capacity . This suggests that 11β-HSD1 plays a role in regulating the magnitude and resolution of inflammatory responses.

When designing experiments using 11β-HSD1 inhibitors or genetic models:

• Consider the timing of intervention (acute vs. chronic inhibition)

• Include appropriate controls to distinguish direct effects from compensatory mechanisms

• Assess both inflammatory cell recruitment and functional parameters (e.g., phagocytic capacity)

• Monitor longitudinal resolution of inflammation rather than single time points

What methodological approaches can distinguish between 11β-HSD1 and 11β-HSD2 activities?

Distinguishing between 11β-HSD1 and 11β-HSD2 activities is crucial in tissues where both enzymes may be expressed. Several approaches can help researchers differentiate between these activities:

The choice of approach depends on whether the researcher is measuring enzymatic activity or protein expression, and whether the experimental system contains both isozymes .

Why might Western blot analysis show multiple bands when detecting 11β-HSD1?

Western blot analysis of 11β-HSD1 can sometimes reveal multiple bands, which may represent:

• The main band at 65-70 kDa represents the glycosylated form of 11β-HSD1

• A band at approximately 36 kDa represents the non-glycosylated form

• Additional bands may represent:

  • Degradation products

  • Differentially glycosylated forms

  • Dimeric or multimeric complexes

  • Non-specific binding

To troubleshoot multiple bands:

• Include appropriate positive controls (liver microsomes are recommended)

• Use protein extraction methods that preserve glycosylation when detecting the native form

• Consider deglycosylation treatments to confirm the identity of higher molecular weight bands

• Validate findings using alternative antibodies targeting different epitopes

What are the critical parameters for successful immunofluorescence detection of 11β-HSD1?

For optimal immunofluorescence detection of 11β-HSD1, researchers should consider:

• Fixation method: As an ER-localized enzyme, aldehyde-based fixatives generally work well

• Permeabilization: Sufficient permeabilization is necessary to access the ER lumen

• Dilution range: 1:200-1:800 is recommended for most applications

• Positive controls: HepG2 cells show reliable expression and can serve as positive controls

• Counterstaining: Consider co-staining with ER markers to confirm subcellular localization

• Blocking: Thorough blocking is essential to minimize background, particularly in tissues with high lipid content

When performing co-localization studies, selecting appropriate markers for the endoplasmic reticulum can provide additional confirmation of specific staining.

How does inhibition of 11β-HSD1 affect glucocorticoid-related side effects?

Recent research has demonstrated that 11β-HSD1 inhibition can mitigate prednisolone-induced adverse effects in clinical settings . This finding has important implications for therapeutic applications and experimental designs involving glucocorticoid treatments. When studying 11β-HSD1 inhibition in combination with glucocorticoid administration, researchers should:

• Include appropriate time-course measurements to capture both immediate and delayed effects

• Monitor multiple physiological parameters beyond the primary endpoint

• Consider tissue-specific responses, as 11β-HSD1 activity varies between tissues

• Account for sex differences in response to both glucocorticoids and 11β-HSD1 inhibition

This emerging research area highlights the potential translational significance of 11β-HSD1 studies and may influence protocol design for both basic and clinical research .