Propargyl-PEG4-thiol

Endonucleases Non-Specific, HL-SAN

OverView

HL-SAN efficiently removes nucleic acids from buffers typically used in protein purification. Due to its high salt tolerance, it is the obvious choice for host-cell DNA removal in settings where salt is added to reduce aggregation. Especially efficient for removing nucleic acids from proteins with high affinity for DNA and RNA. Proven performance during lysis and early stages of protein purification processes, as well as high-salt eluates. Cold-adapted enzyme with excellent performance also at ambient temperatures and during over-night digestion at 4°C.

Recommended applications:

Complete removal of DNA 

Viscosity reduction

Key advantages with this Salt Active Nuclease:

• Non specific endonuclease
• Optimum activity at high salt concentration (0.5 M NaCl)
• Active at low temperatures (20% at 6ºC)
• Easily inactivated
• Broad pH range
• Temperature stable

Figures

Figure 1. Optimum activity in solutions with high salinity

HL-SAN has optimum activity at ∼0.5 M NaCl, but operates at a broad range of [NaCl] and [KCl]. The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5, 5 mM MgCl₂ with varying [NaCl] or [KCl]. The maximum activity was set to 100%.

Figure 2. Temperature and activity

HL-SAN has optimum activity at ~35°C, but works over a broad temperature range (20% activity at 10°C and 50°C). The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5 containing 5 mM MgCl₂ and 0.5 M NaCl.

Fig 3. The effect of MgCl₂ and MnCl₂ concentration on the HL-SAN activity.

The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5, 0.5 M NaCl and with varying concentrations of MgCl₂ or MnCl₂. The activity of the sample containing 5 mM MgCl₂ was set to 100%.

Figure 4. HL-SAN activity vs pH/[NaCl]

The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer with different pHs and different concentrations of NaCl. All buffers contained 5 mM MgCl₂. The nature of the buffer was pH-dependent, but generally the NaCl-optimum was the same in all buffers/pHs. The exception was etanolaminbuffer at pH 9 and pH 9.5 in which the NaCl-optimum was shifted to the left (not shown).

Figure 5. Buffer composition affects substrate preference

Without NaCl, the specificity towards ssDNA and dsDNA is similar. At 0.5 M NaCl, the activity towards dsDNA increases, while the activity towards ssDNA is unaffected.

Figure 6. HL-SAN digests ssDNA to ~5-13 nt, and dsDNA to ~5-7 nt

The size of the end products from ssDNA varies from ~5-13 nt, while dsDNA is digested to around ~5-7 nt. The size of the end products seems to depend on the DNA sequence. Substrates 1 and 2 were ssDNA with different sequences and substrates 3 and 4 were dsDNA with similar sequences but with a FAM-label at different ends. Substrate 5 was dsDNA with the same sequence as substrate 3 and 4 but with a FAM-label at both ends.

‍

Figure 7. HL-SAN activity decreases with increasing concentrations of glycerol

The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5, 5 mM MgCl₂, 0.5 M NaCl and with increasing concentrations of glycerol. The activity of the control not containing glycerol was set to 100%.

Figure 8. The activity of HL-SAN at different concentrations of imidazole

The activity of HL-SAN was tested in a 25 mM Tris-HCl buffer, pH 8.5, 5 mM MgCl₂, 0.5 M NaCl and with varying concentrations of imidazole. The activity of the control not containing imidazole was set to 100%.

HL-SAN efficiently removes nucleic acids from buffers typically used in protein purification. Due to its high salt tolerance, it is the obvious choice for host-cell DNA removal in settings where salt is added to reduce aggregation. Especially efficient for removing nucleic acids from proteins with high affinity for DNA and RNA. Proven performance during lysis and early stages of protein purification processes, as well as high-salt eluates. Cold-adapted enzyme with excellent performance also at ambient temperatures and during over-night digestion at 4°C.

TCO-PEG24-DBCO is a monodisperse, long PEG linker featuring a trans-cyclooctene and a DBCO group. DBCO easily reacts with azides through click chemistry, while the TCO group readily reacts with tetrazine-containing compounds.

TCO-PEG24-DBCO is a monodisperse, long PEG linker featuring a trans-cyclooctene and a DBCO group. DBCO easily reacts with azides through click chemistry, while the TCO group readily reacts with tetrazine-containing compounds.

Description

Specifications

Clone	IHC008
Source	Mouse Monoclonal
Positive Control	Ovarian Carcinoma (Non-Mucinous Carcinoma), Thyroid Carcinoma, Renal Cell Carcinoma
Dilution Range	1:200

PAX-8 is a member of the paired box (PAX) family of transcription factors, which are key regulators in early development. This protein plays a role in development of thyroid follicular cells and the expression of thyroid-specific genes, with mutations in the PAX-8 gene linked to thyroid follicular carcinomas, atypical thyroid adenomas, and thyroid dysgenesis. The PAX-8 protein is expressed in simple ovarian inclusion cysts and non-ciliated mucosal cells of the fallopian tubes, but is absent from normal ovarian surface epithelial cells. PAX-8 is also not expressed in normal lung or lung carcinomas. Reports have associated PAX-8 expression with renal carcinoma, nephroblastoma, and seminoma, and have indicated PAX-8 as a useful marker for renal epithelial tumors, ovarian cancer, and for differential diagnoses in lung and neck tumors. Anti-PAX-8 can be useful in determining the primary site of invasive micropapillary carcinomas of ovary from bladder, lung, and breast, when used in adjunct with a panel of organ-specific markers such as uroplakin, mammaglobin, and TTF-1.